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Cabin Canyon Spur, Clark County, Nevada, United States
Ridge(s) - a long narrow elevation with steep sides, and a more or less continuous crest
Mindat.org Region:
Clark Co., Nevada, USA
Clark County, Nevada, United States
114° 5' 37" W
Lat/Long (dec):
36.64387,-114.09386
Köppen climate type:
BWk : Cold desert climate
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Mindat.org localities near to this feature
Virgin Mountain property, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 37' 47" N 114° 5' 4" W 1.7km (1.1 miles)
Virgin Mtn. Beryllium Deposits (Virgin Mountain Chrysoberyl Property; Mica-Beryl Prospects), Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 37' 47" N 114° 5' 4" W 1.7km (1.1 miles)
Silver Leaf mine (Tri-State Metals mine; Mesquite; Deer Trail; Cabin Canyon), Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 38' 6" N 114° 4' 37" W 1.8km (1.1 miles)
Walker Tri-State Mine (Silver Leaf; Silver Leaf No. 1 Claims; Mesquite; Deer Trail; Cabin Canyon), Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 38' 6" N 114° 4' 37" W 1.8km (1.1 miles)
Santa Cruz Mine (Taglo prospect; Teagle mine; Emerald group; Blue Jay group; Rising Star; Rattle Snake), Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 39' 2" N 114° 3' 24" W 3.4km (2.1 miles)
Blue Beetle No. 1 Claim (Walker Prospect), Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 37' 15" N 114° 7' 37" W 3.9km (2.4 miles)
Blackrock No. 1 Claim, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 39' 29" N 114° 3' 11" W 4.0km (2.5 miles)
Whitney Ridge, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 39' 59" N 114° 3' 2" W 4.6km (2.9 miles)
North Star Mica Prospect, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 36' 21" N 114° 7' 21" W 4.9km (3.1 miles)
Virgin Mountains, Clark Co., Nevada, USA 36° 35' 50" N 114° 5' 44" W 5.2km (3.2 miles)
Great Eastern mine, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 37' 32" N 114° 8' 50" W 5.2km (3.2 miles)
Blue Bell (Virginia Copper No. 1), Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 37' 46" N 114° 9' 26" W 5.9km (3.7 miles)
Hummingbird claims (Hummingbird group; Rockland Mine), Virgin Peak (Bunkerville Mt.), Virgin Mountains, Mohave County, Arizona, USA 36° 39' 51" N 114° 1' 45" W 6.2km (3.9 miles)
Western Beryl prospect (North Star prospect; Rock Candy prospect), Mohave County, Arizona, USA 36° 39' 0" N 114° 1' 0" W 6.9km (4.3 miles)
Key West Mine, Whitney Pocket Basin, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 37' 6" N 114° 9' 55" W 7.0km (4.3 miles)
South Valley No. 4 Claim, Gold Butte Mining District, Clark Co., Nevada, USA 36° 37' 40" N 114° 10' 21" W 7.2km (4.5 miles)
East Nos. 51-57 Claims (Virgin Mountains Isa; East No5. 51-57), Whitney Pocket Basin, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 36' 47" N 114° 10' 4" W 7.4km (4.6 miles)
Mica Queen prospect, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 34' 55" N 114° 3' 6" W 7.8km (4.9 miles)
Lime Kiln Canyon, Mohave County, Arizona, USA 36° 39' 0" N 114° 0' 0" W 8.4km (5.2 miles)
Thanksgiving Lode (East Nos. 1-9 Claims; Virgin Mountains Isa), Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 36' 16" N 114° 11' 8" W 9.3km (5.8 miles)
Cedar Wash, Mohave County, Arizona, USA 36° 35' 48" N 113° 59' 47" W 10.1km (6.3 miles)
Lime Kiln Mine, Jacobs Well, Red Pocket Tanks area, Mohave County, Arizona, USA 36° 37' 37" N 113° 58' 34" W 10.6km (6.6 miles)
Black Diamond Mine (Black Metal Mine; Polaris Mine), Black Diamond Mining District, Mohave County, Arizona, USA 36° 37' 36" N 113° 58' 31" W 10.7km (6.7 miles)
Albert Nay Prospect, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 32' 49" N 114° 6' 37" W 10.9km (6.7 miles)
Virgin Mountain deposit, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 32' 47" N 114° 8' 30" W 11.7km (7.2 miles)
Roadside Prospect (Virgin Mountains Isa), Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 32' 13" N 114° 4' 53" W 11.9km (7.4 miles)
Valley View Mine, Mohave County, Arizona, USA 36° 41' 0" N 113° 58' 0" W 12.2km (7.6 miles)
Ranch View (Whitney Pass Prospect; Virgin Mountains Isa), Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA 36° 32' 0" N 114° 4' 3" W 12.5km (7.8 miles)
Jacob's Ranch, Mohave County, Arizona, USA 36° 31' 59" N 114° 0' 0" W 14.9km (9.2 miles)
Cottonwood Wash, Mohave County, Arizona, USA 36° 42' 0" N 113° 52' 59" W 19.8km (12.3 miles)
Kotulskite, etc.
Key West Mine, Whitney Pocket Basin, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA
Virgin Mountain property, Bunkerville Mining District, Virgin Mountains, Clark Co., Nevada, USA
Kotulskite
Minerals recorded nearby (within 19.8 km)
ⓘ Albite
ⓘ 'Albite-Anorthite Series'
ⓘ Almandine
ⓘ Azurite
ⓘ Beryl
ⓘ 'Biotite'
ⓘ Brochantite
ⓘ Chalcanthite
ⓘ 'Chlorite Group'
ⓘ Chromite
ⓘ Chrysoberyl
ⓘ Chrysocolla
ⓘ Corundum
ⓘ Cuprite
ⓘ Dumortierite
ⓘ Epidote
ⓘ Erythrite
ⓘ 'Fayalite-Forsterite Series'
ⓘ 'Feldspar Group'
ⓘ 'Garnet Group'
ⓘ 'Garnierite'
ⓘ Glaukosphaerite
ⓘ 'Halloysite'
ⓘ Hematite
ⓘ Heterogenite
ⓘ 'Hornblende'
ⓘ 'Hypersthene'
ⓘ Kotulskite
ⓘ Kyanite
ⓘ Magnetite
ⓘ Malachite
ⓘ Merenskyite
ⓘ 'Mica Group'
ⓘ Michenerite
ⓘ Microcline
ⓘ Millerite
ⓘ Molybdenite
ⓘ Moncheite
ⓘ Muscovite
ⓘ var: Sericite
ⓘ Opal
ⓘ Palladium
ⓘ Pentlandite
ⓘ Phenakite
ⓘ Polydymite
ⓘ Pyrrhotite
ⓘ 'Scapolite'
ⓘ Scheelite
ⓘ Schorl
ⓘ Sillimanite
ⓘ Sperrylite
ⓘ Spinel
ⓘ Staurolite
ⓘ Talc
ⓘ 'Tourmaline'
ⓘ Vanadinite
ⓘ Violarite
ⓘ Vysotskite
Rock types recorded nearby (within 19.8 km)
ⓘ Amphibolite
ⓘ Hornblendite
ⓘ 'Mica schist'
ⓘ 'Pegmatite'
Entries shown in red are rocks recorded for this region.
Hen Spring Spring(s) 36° 39' 55" N 114° 5' 52" W 2.4km (1.5 miles)
Cow Canyon Valley 36° 39' 1" N 114° 3' 53" W 2.7km (1.7 miles)
Silver Leaf Mine Mine(s) 36° 37' 54" N 114° 3' 50" W 3.0km (1.8 miles)
Santa Cruz Mine Mine(s) 36° 39' 6" N 114° 3' 35" W 3.2km (2.0 miles)
Virgin Peak Ridge Ridge(s) 36° 36' 48" N 114° 5' 55" W 3.4km (2.1 miles)
Virgin Mountain Natural Area Park 36° 36' 33" N 114° 5' 20" W 3.9km (2.4 miles)
Cabin Canyon Valley 36° 40' 24" N 114° 4' 3" W 4.0km (2.5 miles)
Virgin Peak Mountain 36° 36' 9" N 114° 6' 41" W 4.8km (3.0 miles)
Great Eastern Mine Mine(s) 36° 37' 28" N 114° 8' 45" W 5.1km (3.2 miles)
Virgin Mountains Mountains 36° 35' 50" N 114° 5' 44" W 5.2km (3.2 miles)
North America Plate Tectonic Plate
USA Country
Nevada, USA State
Clark Co., Nevada, USA County
Quick NavTopMineral ListRock TypesFeaturesMindat Regions


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Bishop Announces Measures to Address Potential Spread of COVID-19
(SAN DIEGO, March 4, 2020) – Bishop Robert McElroy, head of the Roman Catholic Diocese of San Diego, issued the following statement today concerning changes parishioners and the public will see at Mass to respond to concerns about the spread of what is now known as COVID-19 or the coronavirus. The diocese is continuing to stay in touch with state and local public health authorities and will be guided their by advice:
"The spread of the new coronavirus into the United States poses a challenge to all of us which demands vigilance, prudence and calm. We know the anxiety level is high, but the level of the virus in our population remains low, far smaller than the seasonal flu.
"Nevertheless, I want to respond to those concerns and examine whether we need to make some temporary changes in our celebration of the Eucharist.
"For now, all parishes will suspend the practice of sharing Communion (the Precious Blood) by cup. All parishes will also take steps toward limiting the distribution of the Eucharist on the tongue. As to the Sign of Peace, that part of the Mass where Catholics traditionally extend hugs or handshakes, people are simply asked to be prudent; touching is not required."
"We remain in close communication with State and County public health authorities and will advise our faith community if further changes are needed."

redpajama_set_name: RedPajamaCommonCrawl


http://bluemondayauctions.com/wp-content/uploads/2017/02/new-website.jpg 598 900 bmAdmin http://bluemondayauctions.com/wp-content/uploads/2017/03/blue-monday-auctions.png bmAdmin2017-02-20 16:36:382017-03-02 10:53:50Welcome to our new website!
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It is extremely refreshing to work with a company whose priority is to raise funds for charity and not just line their own pockets.
Blue Monday have great items, a slick operation and generate fantastic results. The team are a pleasure to work with.
From the outset you were very flexible with arrangements and the feedback from my guests on the variety and quality of lots was brilliant.


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Home/NEWS/ENTERTAINMENT/Celebrity News/Why Tom Brady Ignores Social Media Commentary – E! Online
Why Tom Brady Ignores Social Media Commentary – E! Online
Mason penelope October 18, 2022
You'll want to huddle up for Tom Brady's advice.
Amid rumors he and wife Gisele Bundchen are headed for divorce, the Tampa Bay Buccaneers quarterback explained his approach to social media.
"I think the point is everyone should just be authentic to who they are," the six-time Super Bowl champ said on the Oct. 17 episode of the Sirius XM podcast Let's Go! With Tom Brady, Larry Fitzgerald and Jim Gray. "You know, some people approach it in different ways and it just has to work for them as part of your personality. So I definitely say things, you know, in the moment with me, my style is to just ignore as much as possible."
After all, his followers—and his haters—don't always see every side to the story. "Most of the people in the conversation don't have a full idea of what's really going on," Brady continued. "Certainly in football it's a challenge because you know, there's so many little nuances and details to a very intricate sport in terms of strategy and what you're trying to accomplish. No one really outside of what the group is can really help us anyway."
Amazon's Prime Video will stream first-ever NFL game on Black Friday next year
Man accused of kidnapping estranged wife in Lacey, hiding her in woods, now in custody
Givenchy's Cult Favorite Black Magic Lipstick Is Finally Back in Stock – E! Online
Aaron Carter Missed Rehab Therapy Session Night Before He Was Found Dead
Walmart Early Black Friday Sale: Get a $300 Suitcase Set for $100 & More Finds at Unbeatable Prices – E! Online
Kanye West Could Buy 'White Lives Matter' Rights For $1 Billion, Owners Say
Breaking Down the Wild Week After Elon Musk Bought Twitter for $44 Billion – E! Online


In a week time, I am going to have a technical interview and I was asked to bring an example of my own work to discuss. I am going to show the project I've been working on for the last 1+ years in my spare time. I know it inside out but I am still worried.
Would you have any advice on how can I prepare myself? I am revising and cleaning my code a little bit, what should I pay the most attention to?
They probably wont be looking for perfection, more likely they'll be looking at how you work, your logical approach and ideas.
If you're showing a home project you've been working on, it's perfectly acceptable for parts of your code to be work in progress. If things need to be improved and you don't have time to do it before your interview, then notate what needs to be done and be open about it - Show that you are aware of priorities.
Most of all though, just show a bit of passion and interest!
I'd go further, and say that parts that are work-in-progress or otherwise buggy are important, because you get to talk about them. You get to talk about how if you started from scratch you might have done something differently, and which problems you are prioritising, and why.
If you're showing a home project you've been working on, it's perfectly acceptable for parts of your code to be work in progress. [..] Show that you are aware of priorities.
100% agree, with a special accent on the priorities part. I'd always take a dev that knows how to choose his priorities over one that "perfects everything" (whatever that even means).
That said, I think that you should have at least some part of the code sleek and well tested, just to show that when you eventually put a priority on something you do it well.
Priorities are super important, yes. I have a lot of things/bugs shoveled into the backlog, including tests on many non-critical parts. Otherwise, nothing would be working even at this point.
Give your interviewers a quick run down of your perspective and the problems you wanted to solve with your solution. This makes it more likely that you all start with the same mental model.
Have a good mental model of the different parts of your application and how they work together. Identify important paths through your application that you can talk about (almost like user stories). For example, you probably don't want to spend a lot of time in your utils package, talking about string helpers and so on.
"Why did you use that library/data structure/database/...?"
Even if the answer is just: "lot's of people use it so there's a large community to ask questions to.", as long as it doesn't look like it's the first time you're thinking about that question, you'll probably be good.
In general, I like to think of an application as a complex topic that I've written an essay about. Now the challenge is to present that essay concisely and to the point in 10-15 minutes.
It is very likely that they spot a loop, a part of your code with some uncommon functions or spread operators and ask you to explain what it does and why you choose to go with that.
Glad my comment helped you!
Most important thing is to be confident! You wrote that code and you know it, its just because of time you might forget some things but you will always have an idea.
The next thing in this week time, find out the shortcoming you faced in building that project. The interviewers are always interested in knowing some cool facts and approaches, the way you bend your thought process!
And be confident, you will rock it !
Thank you! :) I think, being confident is the hardest part since I know how many places where the code is crap. Good thing is that it used to be a lot worse and I am improving all the time.
I have a ton of cool stories about this project and various shortcomings, yep.
I know you are worried, but I guarantee working with your own code is going to be so much nicer than if they were asking you to sit down and write it from scratch. Keep in mind they are not looking for perfection, they likely just want to see how you work and think through things.
It actually went really nicely. First, I showed the project in action and explained how it came to be and what the goals are. After that, I was asked to show one component of my choice and I went with a recently refactored one and explained my motivations for the refactor (btw, it was my first try on react hooks!).
Then, on another project where I did backend as well, I showed two parts of the system I've been doing by myself, and we discussed the decisions made, SQL query efficiency and stuff like that.
I think it is definitely a better way for doing technical interviews, compared to coding on the spot.


I know some of us moan about technology and Big Brother watching you all the time. But after today, I am really thankful for it.
My Daughter had an after school club today, so instead of her getting the bus home. We arranged she would text me and I would pick her up.
I had a busy day today, where the day just flew by. I checked my phone to see what the time was and it was coming up to just before 4pm. So I thought I would have a quick coffee before she text me.
This is where I am thankful for technology. We have a tracker on our phones and I can track to see where she is. When she is at school her phone is always turned off. But I do know that when she goes to after school clubs she turns it on. So I thought I would just check while having my coffee. How glad am I that I did.
The tracker said she was half way home, so I checked the bus and it was due in 2 minutes at our stop, (yes I can track the bus as well), the bus came and did not stop and went sailing past. Queue me going into panic mode, I kept ringing her phone no reply, tried face time no reply. Luckily my Mum was with me, so I grabbed her keys and we ran out the door into the car to look for her. I saw her walking up the road. Now I live on the same road as a major attraction, it is very busy, one side has a pavement the other a grass verge. She was walking along on the grass verge. I stopped the car and Mum jumped out (in her slippers) and ran across to her while I turned the car around and picked them up.
I am so glad my Mum got to her first or I would have killed her and then hugged her. Her after school club was cancelled, her phone had broken and she missed the bus as had not been told that the club was cancelled, so as she couldn't contact me she walked home.
Luckily, her Dad has fixed her phone and she knows to walk on the pavement side, but honestly I have never been so scared, it is amazing what goes through your mind.
Today I was so very grateful to modern technology.
Mothergeek has also spoken about when should kids have smartphones. It is every childs dream and every parents nightmare as to when to give in and let your child have a phone.
I was scared, you can't imagine the thoughts going through my head, the feeling of relief when I saw her (albeit the wrong side of the road), was immense.
That must have been terrifying! It just goes to show technology isn't the devil.
I used the Glympse app. It allows you to turn it on for a set amount of time (perfect for a journey) and then you send it to who you want to see. Then again if her phone was broken it wouldn't have been a lot of good.
You must have been terrified! Definitely Thankful for technology. I never thought of the ways it helps us take better care of children. I am one of those people that mostly resents the constant connectivity, the not being able to get away from it no matter where I am, and the creepy big brother aspect, but you are absolutely right, technology does have so many positives and this is one of them. Thanks for sharing!
I lost my daughter at a large park once. I looked down at my baby, looked up and she had disappeared. I called for her and searched and searched. Finally a breathless 20 minutes later I found her on the other side of the park. Losing a child is so scary!
That must have been scary, it is horrible when you suddenly find out that they are not where they are supposed to be, and all sorts go through your mind.


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Posted June 12, 2015 by Fanny Nicolaisen & filed under This Week in South Sudan
Saturday 6 June
The SPLA launched an operation in Western Bahr el Ghazal to find the suspected rebels who killed at least 10 soldiers two weeks ago.
South Sudanese presidency dismissed media reports over President Salva Kiir bad health and alleged speculations that Minister of Defence, Kuol Manyang Juuk would replace him.
According to a government official, South Sudan is set to unveil its' first-ever coins into the market during the fourth independence anniversary on 9 July.
Radio Tamazuj: South Sudan conflict situation overview, 7 June.
Monday 8 June
Abyei Administration Area (AAA) and Western Bahr el Ghazal state signed a Memorandum of Understanding (MoU) concerning institutional co-operation and capacity building for the displaced officials of Abyei region.
Kuol Madang Kuol, the rebel-appointed 'governor' of Northern Bahr al Ghazal claimed that their forces captured an SPLA garrison in Aweil West County on Sunday morning. Government officials later denied this.
South Sudan's military said it controls the oil fields in Unity and Upper Nile States, contrary to rebel claims that they captured them after heavy fighting.
Tuesday 9 June
The SPLA claimed it repulsed an attack allegedly carried out by the SPLA (IO) Panakuac, and took control of several areas previously held by the rival forces. The SPLA (IO) later denied this.
At least three people were shot dead in a random shooting that occurred in Maridi town in Western Equatoria state.
In Western Equatoria state a spokesperson for the Revolutionary Movement for National Salvation (REMNASA), claimed their forces destroyed a production unit belonging to the SPLA 4 km from Maridi town.
Torit County Commissioner said that Hiyala Payam has been deserted by local residents since inter-communal clashes last week when about 149 houses were torched.
Wednesday 10 June
The former caretaker governor of Northern Bahr al Ghazal state misappropriated over 160,000 US dollars' worth of public funds, according to an investigation commissioned by the state legislative assembly.
At least two SPLA soldiers were reportedly killed and 10 others wounded in clashes with the SPLA (IO) rebels in Manyo County Upper Nile state.
Thursday 11 June
According to a SPLA (IO) spokesperson, rebel leader Riek Machar is in South Africa to participate in the African Union (AU) summit, starting next Monday, by heads of state and government in the continent.
The governor of South Sudan's Central Bank, Kornelio Koriom, advised the country's lawmakers against returning the management of letters of credit under the bank's authority.
South Sudan's government and the SPLM-IO have endorsed the proposed inclusion of new actors to the IGAD – led peace process.
International Crisis Group: Disunity State: A journey through South Sudan's Unity state.


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Reading: Volvo Polestar 2: Is It Worth Rivalling the Tesla Model 3?
© 2022 BroncoDrive Company. All Rights Reserved.
> Overview > Volvo Polestar 2: Is It Worth Rivalling the Tesla Model 3?
Volvo Polestar 2: Is It Worth Rivalling the Tesla Model 3?
Pavlo Prannyk
The fully-electric Polestar 2 is the latest addition to Volvo's Polestar brand. The sport sedan is the first fully-electric vehicle from this brand, starting at around $60,000. Although Tesla is a well-known name in electric cars, the Polestar 2 is designed to rival the Tesla Model 3.
The Polestar 2 comes standard with all-wheel drive and offers a range of about 235 miles when fully charged. Car enthusiasts would be impressed with the stats on this car. It features 400 horsepower and about 500 pound-feet of torque. These numbers give you a 0-60 time of about 4.7 seconds!
But the question that many are asking is whether the Volvo Polestar 2 is an excellent rival to the famous Tesla vehicles. In this guide, we will go over some of the exciting features of the Polestar, so you can decide whether it's best for you.
Exciting Features on the Volvo Polestar 2
The Volvo Polestar 2 comes with some interesting quirks and features, with anyone who wants an excellent electric vehicle would be impressed.
Activity Key
If you're familiar with Volvo vehicles, you will notice that the key design is the same as the Polestar, except for the Polestar design on it. But asides from the standard key, you will also be given an activity key, which is much smaller. It's ideal for outdoor activities and adventures that might require you to move around a lot, as you can easily store the activity key. The key is also waterproof up to 30 feet.
Neat Lighting and Design
There are little touches here and there on the Polestar 2 that gives you a neat and attractive vehicle. The taillight does a little dance when you turn on the car, which is a pretty neat trick. You also see a Polestar logo on the roof, which lights up.
Another unique touch is the Polestar logo itself, which you can find everywhere on the car. You can see the logo on the front, back, roof, door sill, and wheel caps. But there's no place on the vehicle that has Polestar written except the brake light. While this doesn't add anything to its performance, it's interesting to note.
Unique Start Feature
When you step into your Volvo Polestar 2, you might be surprised to see that there's no start button. The car recognizes that you're there, and you can shift into drive and be on your way. And when you want to stop the car, you park it and leave it, and it turns off on its own. The gear lever has an unorthodox design with a hexagonal shape and a Polestar logo illuminated in the center.
Infotainment and Technology
Technology is usually a big part of electric vehicles, and since the Polestar 2 is competing with the Tesla Model 3, you can expect some impressive tech features. The dashboard's center is a large, portrait-designed screen that sets the Polestar apart from previous models.
The infotainment screen will remind you of a large tablet and includes four sections: navigation, media player, connect your phone, and a virtual assistant. Navigation is done with Google Maps for ease. The screen also has climate controls at the bottom.
It is sensitive to touch and easy to navigate. It also provides different opportunities for customization, to design how your climate controls work, and more. There is a 360-camera that shows your car from a bird's eye view and makes it easy to navigate tight spaces and park. It also comes with other features for driver-assist and checking your cars for errors.
Gauge Cluster
Like in many modern cars, the gauge cluster on the Polestar 2 is designed as a full screen. It shows you detailed information like the speed limit for the road you're currently on, your gear selection, current speed, battery life, current mileage, and the charge vs. power. You can also put the navigation map and trip odometer to the gauge cluster.
Although not as attractive as in Tesla, the Volvo Polestar 2 has a simple and minimalistic interior design. There are also neat trims like aluminum trim on the volume button. There is a cup holder outside and inside the center console. The vehicle comes with a cloth for cleaning the infotainment system.
The glove box is large and well-designed as a shelf. And the center climate vents are on top of the dashboard, which is a strange placement. The backseat is a bit tight, but that's because the Polestar 2 isn't a large vehicle. The seatbelts are also yellow.
You notice the sunroof with no crossbar or anything interrupting it when you look up. It gives you an open feeling while in your car. But, it doesn't open.
The cargo area in the Volvo Polestar 2 is designed as a hatchback, so the entire back lifts. This gives you more space than sedan vehicles usually have. There are also different features to maximize your storage space.
You can set up a divider in the middle of the cargo area to separate stuff or stop them from crashing into each other. There is also a lower cargo area for extra storage and a net on the right side of the area where you can put small things that you don't want rolling around.
There is also a cargo area in the front of the car since there is no engine.
The Polestar 2 has a gorgeous exterior styling as a sedan vehicle, so you can be sure to drive with style wherever you go. You can also adjust the side mirrors easily, and the entire mirror would move when you do so. The model is slightly higher than a regular sedan, with all-wheel drive and a capable exterior for rough driving, making it a bit closer to an SUV crossover.
Polestar 2 vs. Model 3
The closest rival to the Polestar 2 is the Tesla Model 3, but the latter is about $5,000 cheaper and offers about a 315-mile range and goes 0-60 in 3.1 seconds. So, Tesla provides more in terms of performance and affordability.
Despite this, the Polestar 2 comes with some fantastic features, impressive technology, and an attractive design that would enhance your driving experience wherever you go.
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Pavlo Prannyk March 15, 2022
Posted by Pavlo Prannyk
Cars were always a big part of my life. From my early childhood, I was dreaming about cars. Now in my adultness, I still have a big interest in cars, and in my free time, I'm sharing my car experience here on this blog. Hope you enjoy it.
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Banking reform was also vigorously promoted in the wake of both slumps. In 1933, the US Glass-Steagall Act stopped commercial bank proprietors from speculating with their customers' deposits. After 2009, the US Dodd-Frank legislation, the Vickers Report in the United Kingdom, and the European Union's Liikanen report similarly aimed to make the banking system more "resilient" to "shocks." Although the shocks to banking were the effect, not the cause, of shocks to the economy, this was ignored: Restore money and banking to order, the argument went, and shocks would stop.
This is the context in which the rise of cryptocurrencies – the latest explosion of monetary mania – should be understood. These "peer-to-peer electronic cash systems" aim to cure economic ills by monetary measures, but this time by bypassing banks altogether. Why do we need these diseased intermediaries, cryptocurrencies' inventors ask, when we can create electronic storage and transaction systems, secure to their users and invisible to would-be controllers?
The technical details of the new cash-generation systems are difficult to grasp; their inspiration is not. The start of Bitcoin in January 2009 coincided with the banking crisis. Banks went bust or were rescued from insolvency by taxpayers. Understandably people wanted to find a way to keep their money and monetary transactions out of the reach of failing banks and voracious tax authorities. The new cryptocurrency offered a solution.
Bitcoin can be seen as an attempt to use new technology to stop money from going bad. For example, the total supply of bitcoins is fixed, as it would be – more or less – for a gold-backed currency. Paradoxically, although it is created out of nothing, it will offer no possibility of money "creation." Bitcoin will be "mined" in diminishing quantities until it is exhausted in 2040, having delivered 21 million digital coins. In other words, there is no elasticity in the currency. This means that long before the mine is exhausted, the currency will run into the same problem as the gold standard: not providing enough money to support a growing economy and population. This would be exacerbated by any tendency to hoard bitcoins.
At the same time, cryptocurrencies provide no security against inflation. Hayek thought that a competitive currency system would eventually lead to a monopoly of the one that kept its value best. But we have, of course, been through exactly this process of weeding out inflationary currencies throughout history, and we ended up with central banks. It is amazing that anyone should consider it necessary to retrace these steps, only to end up in the same place.
The fact is that human societies have discovered no better way to keep the value of money roughly constant than by relying on central banks to exercise control over its issue and to act directly or indirectly on the volume of credit created by the commercial banking system. The Hayekian diagnosis of the last crisis – excessive creation of credit by the banks – is correct as far as it goes. But one has only to ask why this happened to understand that there are no mechanical answers to the question or solutions to the problem. It's not quite true to say, "Look after the economy and money will look after itself." But it is nearer the truth than the belief that monetary reform on its own will cure the problems of a sick economy.


Property Location Located in Isle of Skye (Kyleakin), Mackinnon Country House Hotel is convenient to Caisteal Maol and Skye Bridge. This hotel is within the vicinity of Plockton Harbour and Skye Serpentarium. Rooms Stay in one of 16 guestrooms featuring flat-screen televisions. Complimentary wireless Internet access keeps you connected, and cable programming is available for your entertainment. Private bathrooms with bathtubs or showers feature complimentary toiletries and hair dryers. Housekeeping is provided daily, and both cribs/infant beds (complimentary) and rollaway/extra beds (surcharge) are available on request. Amenities Take in the views from a terrace and a garden and make use of amenities such as complimentary wireless Internet access. Dining Enjoy a satisfying meal at a restaurant serving guests of Mackinnon Country House Hotel. Quench your thirst with your favorite drink at a bar/lounge. A complimentary full breakfast is served daily from 8 AM to 9:30 AM. Business, Other Amenities The front desk is staffed during limited hours. Free self parking is available onsite.


text:
Q: In Java, it's possible discover what was the current method/methods executed? I'm working with threads but after a time, most of them stop doing their job. The first thing I thought was a deadlock, but all are with state RUNNING.
I suppose there is an error in my logic or a new characteristic that I not realized and I must handle (it's a webcrawler).
Is it possible to get the current executing method or operation? I want this to see where my threads are trapped.
EDIT: I think that is something I need to handle or there is error in my logic because this happens after a time executing, not imeddiatly after the start.

A: You can get the current stack trace in Java.  You will get an array of StackTraceElement elements.
The first item in the array is the currently executing method.
See the following question for how to get the stack trace:

Get current stack trace in Java

Code might look like:
StackTraceElement[] trace = Thread.currentThread().getStackTrace();
StackTraceElement yourMethod = trace[1];
System.out.println(yourMethod.getMethodName());

A: A debugger is the way to go. This is what they are designed for.
Java debuggers with threading support are built into both the Eclipse and Netbeans IDEs.

A: Make VM to dump the threads (Ctrl-Break). Find your threads in the list. Look at the topmost stacktrace method. Done.

A: You have 2 options:

*

*Use debug to get some understanding that was executed and what not.

*Use a lot of logmessages (you can also produce stacktraces in that messages)

A: Thread dumps are the right solution for the problem. If you want to do it programmatically within the process (some kind of monitoring logic), then java.lang.management.ThreadMXBean provides access to all threads along with their current stacks at the time.

A: It is, throw an exception, catch it immediately and save the stack. This is about as performant as asking an elephant to fly overseas but it's possible since it sort of extracts the current call stack to something you can work with.
However, are you sure you haven't run into a livelock?

A: Do you suppose your web crawler program is in a loop processing the same urls. Add some high level logging so each thread writes what it's processing.

redpajama_set_name: RedPajamaStackExchange


text:
<?xml version="1.0" encoding="utf-8"?>
<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN"
    "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">

    <title>Date</title>
    <meta http-equiv="Content-Type" content="text/html; charset=UTF-8" />
    <link rel="stylesheet" href="../css/reset.css" type="text/css" media="screen" />
<link rel="stylesheet" href="../css/main.css" type="text/css" media="screen" />
<link rel="stylesheet" href="../css/github.css" type="text/css" media="screen" />

            Ruby on Rails 4.2.1

            Class
            Date

                <

                    Object

/Users/tony/.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/acts_like.rb

/Users/tony/.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb

/Users/tony/.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/conversions.rb

/Users/tony/.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/zones.rb

/Users/tony/.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/object/json.rb

Methods

#

                <=>

A

                acts_like_date?,

                advance,

                ago,

                at_beginning_of_day,

                at_end_of_day,

                at_midday,

                at_middle_of_day,

                at_midnight,

                at_noon

B

                beginning_of_day,

                beginning_of_week,

                beginning_of_week=

C

                change,

                compare_with_coercion,

                compare_without_coercion,

                current

D

                default_inspect

E

                end_of_day

F

                find_beginning_of_week!

I

                in,

                inspect

M

                midday,

                middle_of_day,

                midnight

N

                noon

R

                readable_inspect

S

                since

T

                to_default_s,

                to_formatted_s,

                to_s,

                to_time,

                tomorrow

X

                xmlschema

Y

                yesterday

Included Modules

              DateAndTime::Calculations

              DateAndTime::Zones

Constants

            DATE_FORMATS
            =
            {
:short        => '%e %b',
:long         => '%B %e, %Y',
:db           => '%Y-%m-%d',
:number       => '%Y%m%d',
:long_ordinal => lambda { |date|
day_format = ActiveSupport::Inflector.ordinalize(date.day)
date.strftime("%B #{day_format}, %Y") # => "April 25th, 2007"
},
:rfc822       => '%e %b %Y',
:iso8601      => lambda { |date| date.iso8601 }
}

Attributes

              [RW]

            beginning_of_week_default

Class Public methods

              beginning_of_week()

            Link

Returns the week start (e.g. :monday) for the current request, if this has
been set (via ::beginning_of_week=).
If Date.beginning_of_week has not been set for the current
request, returns the week start specified in
config.beginning_of_week. If no config.beginning_of_week was
specified, returns :monday.

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 17
def beginning_of_week
  Thread.current[:beginning_of_week] || beginning_of_week_default || :monday
end

              beginning_of_week=(week_start)

            Link

Sets Date.beginning_of_week to a week start (e.g. :monday) for
current request/thread.

This method accepts any of the following day symbols: :monday, :tuesday,
:wednesday, :thursday, :friday, :saturday, :sunday

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 25
def beginning_of_week=(week_start)
  Thread.current[:beginning_of_week] = find_beginning_of_week!(week_start)
end

              current()

            Link

Returns Time.zone.today when
Time.zone or config.time_zone are set, otherwise
just returns Date.today.

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 46
def current
  ::Time.zone ? ::Time.zone.today : ::Date.today
end

              find_beginning_of_week!(week_start)

            Link

Returns week start day symbol (e.g. :monday), or raises an ArgumentError
for invalid day symbol.

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 30
def find_beginning_of_week!(week_start)
  raise ArgumentError, "Invalid beginning of week: #{week_start}" unless ::Date::DAYS_INTO_WEEK.key?(week_start)
  week_start
end

              tomorrow()

            Link

Returns a new Date representing the date 1 day
after today (i.e. tomorrow's date).

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 41
def tomorrow
  ::Date.current.tomorrow
end

              yesterday()

            Link

Returns a new Date representing the date 1 day ago
(i.e. yesterday's date).

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 36
def yesterday
  ::Date.current.yesterday
end

Instance Public methods

              <=>(other)

            Link

              Also aliased as: compare_without_coercion

              Alias for: compare_with_coercion

              acts_like_date?()

            Link

Duck-types as a Date-like class. See Object#acts_like?.

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/acts_like.rb, line 5
def acts_like_date?
  true
end

              advance(options)

            Link

Provides precise Date calculations for years,
months, and days. The options parameter takes a hash with any
of these keys: :years, :months,
:weeks, :days.

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 110
def advance(options)
  options = options.dup
  d = self
  d = d >> options.delete(:years) * 12 if options[:years]
  d = d >> options.delete(:months)     if options[:months]
  d = d +  options.delete(:weeks) * 7  if options[:weeks]
  d = d +  options.delete(:days)       if options[:days]
  d
end

              ago(seconds)

            Link

Converts Date to a Time (or
DateTime if necessary) with the time portion
set to the beginning of the day (0:00) and then subtracts the specified
number of seconds.

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 53
def ago(seconds)
  in_time_zone.since(-seconds)
end

              at_beginning_of_day()

            Link

              Alias for: beginning_of_day

              at_end_of_day()

            Link

              Alias for: end_of_day

              at_midday()

            Link

              Alias for: middle_of_day

              at_middle_of_day()

            Link

              Alias for: middle_of_day

              at_midnight()

            Link

              Alias for: beginning_of_day

              at_noon()

            Link

              Alias for: middle_of_day

              beginning_of_day()

            Link

Converts Date to a Time (or
DateTime if necessary) with the time portion
set to the beginning of the day (0:00)

              Also aliased as: midnight, at_midnight, at_beginning_of_day

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 65
def beginning_of_day
  in_time_zone
end

              change(options)

            Link

Returns a new Date where one or more of the
elements have been changed according to the options parameter.
The options parameter is a hash with a combination of these
keys: :year, :month, :day.

Date.new(2007, 5, 12).change(day: 1)               # => Date.new(2007, 5, 1)
Date.new(2007, 5, 12).change(year: 2005, month: 1) # => Date.new(2005, 1, 12)

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 125
def change(options)
  ::Date.new(
    options.fetch(:year, year),
    options.fetch(:month, month),
    options.fetch(:day, day)
  )
end

              compare_with_coercion(other)

            Link

Allow Date to be compared with Time by converting to DateTime and relying on the <=> from there.

              Also aliased as: <=>

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 134
def compare_with_coercion(other)
  if other.is_a?(Time)
    self.to_datetime <=> other
  else
    compare_without_coercion(other)
  end
end

              compare_without_coercion(other)

            Link

              Alias for: <=>

              default_inspect()

            Link

              Alias for: inspect

              end_of_day()

            Link

Converts Date to a Time (or
DateTime if necessary) with the time portion
set to the end of the day (23:59:59)

              Also aliased as: at_end_of_day

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 83
def end_of_day
  in_time_zone.end_of_day
end

              in(seconds)

            Link

              Alias for: since

              inspect()

            Link

              Also aliased as: default_inspect

              Alias for: readable_inspect

              midday()

            Link

              Alias for: middle_of_day

              middle_of_day()

            Link

Converts Date to a Time (or
DateTime if necessary) with the time portion
set to the middle of the day (12:00)

              Also aliased as: midday, noon, at_midday, at_noon, at_middle_of_day

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 73
def middle_of_day
  in_time_zone.middle_of_day
end

              midnight()

            Link

              Alias for: beginning_of_day

              noon()

            Link

              Alias for: middle_of_day

              readable_inspect()

            Link

Overrides the default inspect method with a human readable one, e.g., "Mon,
21 Feb 2005"

              Also aliased as: inspect

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/conversions.rb, line 66
def readable_inspect
  strftime('%a, %d %b %Y')
end

              since(seconds)

            Link

Converts Date to a Time (or
DateTime if necessary) with the time portion
set to the beginning of the day (0:00) and then adds the specified number
of seconds

              Also aliased as: in

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/calculations.rb, line 59
def since(seconds)
  in_time_zone.since(seconds)
end

              to_default_s(format = :default)

            Link

              Alias for: to_s

              to_formatted_s(format = :default)

            Link

Convert to a formatted string. See DATE_FORMATS for predefined formats.

This method is aliased to to_s.

date = Date.new(2007, 11, 10)       # => Sat, 10 Nov 2007

date.to_formatted_s(:db)            # => "2007-11-10"
date.to_s(:db)                      # => "2007-11-10"

date.to_formatted_s(:short)         # => "10 Nov"
date.to_formatted_s(:long)          # => "November 10, 2007"
date.to_formatted_s(:long_ordinal)  # => "November 10th, 2007"
date.to_formatted_s(:rfc822)        # => "10 Nov 2007"
date.to_formatted_s(:iso8601)       # => "2007-11-10"

Adding your own date formats to #to_formatted_s

You can add your own formats to the Date::DATE_FORMATS hash. Use the format
name as the hash key and either a strftime string or Proc instance that
takes a date argument as the value.

# config/initializers/date_formats.rb
Date::DATE_FORMATS[:month_and_year] = '%B %Y'
Date::DATE_FORMATS[:short_ordinal] = ->(date) { date.strftime("%B #{date.day.ordinalize}") }

              Also aliased as: to_s

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/conversions.rb, line 51
def to_formatted_s(format = :default)
  if formatter = DATE_FORMATS[format]
    if formatter.respond_to?(:call)
      formatter.call(self).to_s
    else
      strftime(formatter)
    end
  else
    to_default_s
  end
end

              to_s(format = :default)

            Link

              Also aliased as: to_default_s

              Alias for: to_formatted_s

              to_time(form = :local)

            Link

Converts a Date instance to a Time, where the time is set to the beginning of the
day. The timezone can be either :local or :utc (default :local).

date = Date.new(2007, 11, 10)  # => Sat, 10 Nov 2007

date.to_time                   # => Sat Nov 10 00:00:00 0800 2007
date.to_time(:local)           # => Sat Nov 10 00:00:00 0800 2007

date.to_time(:utc)             # => Sat Nov 10 00:00:00 UTC 2007

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/conversions.rb, line 81
def to_time(form = :local)
  ::Time.send(form, year, month, day)
end

              xmlschema()

            Link

                Source:
                show

# File ../../../.rbenv/versions/2.2.1/lib/ruby/gems/2.2.0/gems/activesupport-4.2.1/lib/active_support/core_ext/date/conversions.rb, line 85
def xmlschema
  in_time_zone.xmlschema
end

redpajama_set_name: RedPajamaGithub


The BG Group reports discovery of gas on block 64/11 offshore China. The Lingshui 22-1-1 exploration well encountered gas-bearing sands in Qiongdongnan basin of the South China Sea.
The well is 130 km (81 mi) offshore in 1,338 m (4,390 ft) water depth south of Hainan Island.
BG Group is the operator of block 64/11 with a 100% working interest. In the event that a gas or oil field on the acreage receives development approval, CNOOC has the right to acquire an interest of up to 51% and would contribute to the cost of development.


text:
FIDIS (Future of Identity in the Information Society [FoIitIS]) (deutsch: Zukunft der Identität in der Informationsgesellschaft) war ein fünfjähriges Projekt im Rahmen des 6. Forschungsrahmenprogramms. Es dauerte von 2004 bis 2009.

Ziel
Das Ziel war, Veränderungen zu untersuchen, denen der Begriff Identität in der sich dynamisch entwickelnden europäischen Informationsgesellschaft unterliegt und dabei Ansätze und Wege zu identifizieren, die eine möglichst reibungsfreie Europäische Integration erreichen. Das multinationale und multidisziplinäre Netzwerk sollte ein besseres Verständnis dafür entwickeln, welchen Beitrag Systeme für ein Identitätsmanagement für die faire Gestaltung der europäischen Informationsgesellschaft leisten können. Hierbei galt es, nationale Unterschiede zu berücksichtigen. So ist etwa der Personalausweis für Erwachsene in Deutschland Pflicht, in dem Vereinigten Königreich bislang jedoch nicht üblich. Ziel des FIDIS war es, die europäische Forschung hinsichtlich von Identitäts- und Identifizierungstechnologien zu integrieren. Das Projekt beschäftigte sich mit den damit verbundenen Sicherheits- und Datenschutzaspekten. Außerdem erarbeiten die Experten in diesem Netzwerk rechtliche, sozioökonomische sowie anwendungsbezogene Anforderungen in Zusammenhang mit Identität und Identifizierung.

Finanzierung, Projektlaufzeit
Die Europäische Union förderte FIDIS im Rahmen des sechsten Europäischen Forschungsprogramms Technologien für die Informationsgesellschaft. Das Projekt lief seit April 2004 für eine Dauer von fünf Jahren und hatte eine Gesamtförderung von 6,1 Millionen Euro. Die Leitung dieses NoE hatte in Deutschland die Johann Wolfgang Goethe-Universität Frankfurt am Main.

Partner im FIDIS Konsortium
Es waren 24 Partner aus Europa an diesem multidisziplinären und multinationalen Projekt beteiligt.
Johann Wolfgang Goethe-Universität (Frankfurt am Main) – Konsortialführung
Gemeinsame Forschungsstelle (JRC), Sevilla
Vrije Universiteit Brussel
Unabhängiges Landeszentrum für Datenschutz Schleswig-Holstein
Institut Europeen D'Administration Des Affaires (INSEAD)
Universität Reading
Universität Tilburg
Katholieke Universiteit Leuven
Universität Karlstad
Technische Universität Berlin
Technische Universität Dresden
Albert-Ludwigs-Universität Freiburg
Masaryk-Universität (Brünn)
VaF Bratislava
London School of Economics and Political Science (LSE)
Technische und Wirtschaftswissenschaftliche Universität Budapest
International Business Machines Corporation
Institut de recherche criminelle de la Gendarmerie Nationale
Netherlands Forensic Institute
Virtual Identity and Privacy Research Center
Europäisches Microsoft Innovations Center
Institute of Communication and Computer Systems
Sirrix
Nomen Nominandum

Forschungsbereiche
Zu den Kerngebieten, die innerhalb des FIDIS Netzwerks erforscht wurden, gehörten:
 Identity of Identity: Definition einer Terminologie zum Thema ID-Management
 Profiling: Implikationen aus der Sammlung von Daten
 Interoperability of IDs and ID management systems
 Forensic Implications: Forensische Implikationen
 De-Identification: Identitätsdiebstahl und Pseudonymität
 HighTech ID
 Mobility and Identity: ID-Management im Bereich Mobilkommunikation

Weblinks
 Home: Future of IDentity in the Information Society
 Deliverables: Future of IDentity in the Information Society
 ISTweb - IST Projects
 Digital Single Market | Digital Economy & Society

Forschungsprojekt in FP6
Sicherheitspolitik der Europäischen Union
Identifikationstechnik
Abkürzung

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A reverse-genetics approach to understanding gene families associated with human disease is presented, using mouse as a model system, and the subsequent mouse phenotyping schedule is described. Because mice defective in a gene of interest, HtrA2, manifested Parkinsonian symptoms, the phenotyping regimen is focused on identifying neurological defects.
Patterson, V. L., Thompson, B. S., Cherry, C., Wang, S. b., Chen, B., Hoh, J. A Phenotyping Regimen for Genetically Modified Mice Used to Study Genes Implicated in Human Diseases of Aging. J. Vis. Exp. (113), e54136, doi:10.3791/54136 (2016).
Age-related diseases are becoming increasingly prevalent and the burden continues to grow as our population ages. Effective treatments are necessary to lessen the impact of debilitating conditions but remain elusive in many cases. Only by understanding the causes and pathology of diseases associated with aging, can scientists begin to identify potential therapeutic targets and develop strategies for intervention. The most common age-related conditions are neurodegenerative disorders such as Parkinson's disease and blindness. Age-related macular degeneration (AMD) is the leading cause of blindness in the elderly. Genome wide association studies have previously identified loci that are associated with increased susceptibility to this disease and identified two regions of interest: complement factor H (CFH) and the 10q26 locus, where the age-related maculopathy susceptibility 2 (ARMS2) and high-temperature requirement factor A1 (HtrA1) genes are located. CFH acts as a negative regulator of the alternative pathway (AP) of the complement system while HtrA1 is an extracellular serine protease. ARMS2 is located upstream of HtrA1 in the primate genome, although the gene is absent in mice. To study the effects of these genes, humanized knock-in mouse lines of Cfh and ARMS2, knockouts of Cfh, HtrA1, HtrA2, HtrA3 and HtrA4 as well as a conditional neural deletion of HtrA2 were generated. Of all the genetically engineered mice produced only mice lacking HtrA2, either systemically or in neural tissues, displayed clear phenotypes. In order to examine these mice thoroughly and systematically, an initial phenotyping schedule was established, consisting of a series of tests related to two main diseases of interest: AMD and Parkinson's. Genetically modified mice can be subjected to appropriate experiments to identify phenotypes that may be related to the associated diseases in humans. A phenotyping regimen with a mitochondrial focus is presented here alongside representative results from the tests of interest.
Age-associated diseases are becoming increasingly prevalent in modern society. As medical science improves and life expectancy increases, the population continues to age and the burden of these diseases grows. Effective treatments are necessary to lessen the impact of debilitating conditions but remain elusive in many cases. Only by understanding the causes and pathology of diseases associated with aging can scientists begin to identify potential therapeutic targets and develop strategies for intervention. Common age-related conditions include neurodegenerative disorders such as Parkinson's disease (PD) and age-related macular degeneration (AMD). PD is the most common movement disorder caused by neurodegeneration in humans. Most PD patients show symptoms such as resting tremor, bradykinesia and rigidity after 50 years of age. Early onset has also been observed in approximately 10% of cases.
AMD is the leading cause of blindness in the elderly, progressively damaging photoreceptors and the retinal pigment epithelium (RPE) in the eye. Central vision is impaired but the peripheral vision is generally unaffected. There are two forms of AMD. In the "dry" form, extracellular protein deposits known as drusen form between the RPE and Bruch's membrane (BM), leading to geographic atrophy and blurring of central vision. The more severe "wet" form results from neovascularization from the choroid across BM into the RPE and photoreceptor layers and can lead to hamorrhaging beneath the retina that causes permanent damage to retinal tissue. Genome wide association studies have previously identified loci that are associated with increased susceptibility to this disease and identified two regions of interest: complement factor H (CFH) on chromosome 1 and the 10q26 locus, where the age-related maculopathy susceptibility 2 (ARMS2) and high-temperature requirement factor A1 (HtrA1) genes are located1-5. Combinations of these alleles increase the likelihood of AMD in a dose dependent manner and specific SNPs can be preferentially associated with either the wet or dry forms of AMD3-6.
CFH acts as a negative regulator of the alternative pathway (AP) of the complement system by inhibiting the activation of C3. A single nucleotide polymorphism (SNP) has been linked to increased risk of AMD, causing exchange of tyrosine 402 in exon 9 with histidine due to a T to C substitution1. In AMD it is believed that the AP is misregulated because of a loss of function of CFH but whether the SNP plays a causal role is unclear. One hypothesis is that the positively charged histidine is thought to negate the ability of CFH to bind to interacting proteins C-reactive protein and heparin sulfate1,7. In vitro studies of CFH Y402H provide conflicting results over functional differences between the variants, and in vivo work in Cfh-/- mice expressing humanized CFH is ongoing8. ARMS2 is located upstream of HtrA1 in the primate genome, although the gene is absent in mice. HtrA1 is a serine protease but ARMS2 is poorly characterized. The linkage disequilibrium between SNPs in the AMD-associated locus has made it difficult to determine the contributions to risk of individual mutations of the genes in this region, but recent work has suggested that it is overexpression of HtrA1 rather than ARMS2 that leads to neovascularization and subretinal protein deposits9-11. However, the close proximity of the genes in this locus may allow for interactions that cannot be studied using randomly inserted transgenes.
In addition to AMD, the HtrA family of serine proteases has been associated with many human diseases. All HtrA proteins contain a serine protease domain followed by at least one C-terminal PDZ domain. HtrA1, HtrA3 and HtrA4 share the greatest homology, consisting of a signal peptide, insulin-like growth factor binding domain, a Kazal protease inhibitor domain, the serine protease domain and a PDZ domain. HtrA2 has a different N-terminus, composed of a mitochondrial localization sequence, transmembrane domain and inhibitor of apoptosis binding domain followed by the protease and PDZ domains12-16. Mammalian HtrA1 is regulated by substrate-induced remodeling in the active site of its protease domain17-20, and HtrA2 can also be modulated by interaction between the serine protease and PDZ domains that suppresses protease activity21. Interestingly, the PDZ domain does not appear to confer similar regulation to HtrA316. The HtrA proteases can also be regulated by extrinsic factors: it was recently demonstrated that there exists a regulatory interaction between HtrA1 and protoporphyrins22 and HtrA2 can be regulated by phosphorylation upon activation of the p38 MAP kinase pathway in a PINK1-dependent manner23. The deletion of individual members of the HtrA family in mice has been documented, however mechanistic effects mostly are unclear partly due to lack of visible phenotypes.
HtrA1 plays an important function in protein quality control and its misregulation or mutation has been associated with many different human diseases including arthritis, cancer and an increased risk of AMD3,4,24-32. Loss of HtrA2 function in neural tissues has been associated with PD phenotypes in humans and mice, while its loss from non-neural tissues results in accelerated aging33-37. HtrA3 dysregulation has been associated with diseases including preeclampsia and certain types of cancer38,39. Up-regulation of HtrA4 has been observed in the placentas of preeclampsia patients but knockout mice do not display an overt phenotype40,41. The lack of phenotypes observed in some knockout mice has been postulated to be a result of compensation between the HtrA family member: it is thought that both HtrA4 and HtrA1 interact with the TGF-B family of proteins, allowing for compensation by HtrA1 upon deletion of HtrA441. Similarly, it is thought that since HtrA1 and HtrA3 have a high degree of domain homologies they might have complementary functions42. However, it has been suggested that HtrA proteins may have partially antagonistic roles, competing to regulate common targets43.
To further investigate these risk factors three humanized knock-in mouse lines were generated. In Cfhtm1(CFH*9)jhoh and Cfhtm2(CFH*9)jhoh, exon 9 of the Cfh gene is replaced with exon 9 of the human homologue. Cfhtm1(CFH*9)jhoh encodes the non-disease associated tyrosine residue at position 402, whereas Cfhtm2(CFH*9)jhoh carries the Y402H, risk-associated SNP. In ARMS2tm1jhoh the human ARMS2 sequence was targeted to a region upstream of HtrA1. A loxP-flanked STOP sequence placed upstream of the gene sequence but downstream of the included UbiC promoter was excised by crossing to OzCre mice, which express Cre recombinase under the control of the Rosa26 promoter, as previously described34. In addition to these knock-in lines, conditional knockout alleles for Cfh and HtrA1 (Cfhtm1jhoh and HtrA1tm1jhoh), as well as the other known HtrA family members were generated: HtrA2 (HtrA2tm1jhoh), HtrA3 (HtrA3tm1jhoh) and HtrA4 (HtrA4tm1jhoh). The germ-line knockouts were created by crossing OzCre mice to animals engineered to flank specific exons with loxP sites, such that deletion causes a frame shift and/or deletion of the active domain (Cfh; exon 3, HtrA1; exons 2-3, HtrA2: exons 2-4, HtrA3; exon 3, HtrA4; exons 4-6)34,41. A neural deletion of HtrA2, deleted using Cre recombinase under control of the Nestin promoter (HtrA2flox;Tg(Nes-cre)1Kln/J), has also been described34. Only mice lacking HtrA2, either systemically or in neural tissues, displayed clear phenotypes, presenting with Parkinsonian phenotypes.
Since some of these genes of interest are posited to be localized to mitochondria11,44-47, and deletion of HtrA2 generated Parkinsonian phenotypes, a phenotyping regimen with a mitochondrial and neurological focus is described here and representative results from the tests of interest are provided. In order to examine genetically engineered mice produced to investigate human, age-related disease thoroughly and systematically an initial phenotyping schedule was established, consisting of a series of tests related to the two main diseases of interest: AMD and Parkinson's.
Ethics statement: Studies involving animals were conducted in compliance with the National Institutes of Health recommendations in the Guide for the Care and Use of Laboratory Animals and Institutional Animal Care and Use Committee (IACUC) at Yale University.
Note: All mice should be subjected to the same testing regimen to limit differences in habituation to handling. Tests should be performed at the same time of day each time.
Note: The hind limb test is conducted daily from postnatal day (P) 4 to P10 to evaluate proximal hind limb muscle strength, weakness and fatigue.
Transport mice to testing area and allow to rest for 30 min before testing.
Prepare testing apparatus by cleaning a 50 ml conical tube with 70% ethanol, pushing two (P4-P8) or one (P9-P10) cotton balls to the bottom of the tube and securing in a vertical position.
Remove one pup from the cage and record weight. Keep pup in holding tray on heat pad whenever not testing.
Gently suspend the hind limbs of the neonate over the rim of the conical tube, such that it is facing down towards the cotton balls. Count the number of pull attempts made and measure latency to fall using a stopwatch.
Repeat once more then label the pup with marker. Afterwards, identify P6 pups by toe clipping. Rub pups gently with bedding from home cage before replacing.
Wipe conical tube with 70% ethanol.
Repeat testing on next pup until the whole litter has been tested.
Dispose of cotton ball and conical tube. Clean all equipment with 70% ethanol.
Note: The weanling observation test is administered daily from P19 to P21 to score movement and general activity levels.
Prepare Plexiglas testing box marked with a 6x6 grid of 2 inch squares on the base by cleaning with 70% ethanol. Set up video equipment to the side such that the whole box is in the viewing field. Keep the set up and surroundings the same each day to avoid introducing novelty.
Remove one weanling from the cage and place in a clean holding cup. Record weight.
Begin video recording by filming the test identification card, with mouse identification number, age and date of testing. Transfer mouse from holding cup to the center square. Time for three minutes, counting the number of lines traversed by both front paws of the mouse during the test.
Return mouse to a clean cage and end video recording.
Clean the testing apparatus with 70% ethanol.
Repeat until all litter is tested, then return all pups to the home cage.
Clean all testing equipment with 70% ethanol.
Using the videos, quantitate the number of rearing events by tallying after visual observation, where a mouse stands with both front paws clear of support, and grooming events, where both front paws are engaged in cleaning.
Note: The grip strength test is conducted on alternate days between P22 and P26, to assess neuromuscular strength.
Transport mice to testing area and allow to rest in home cage for 30 min before testing.
Prepare apparatus by cleaning a Plexiglass box and mesh grid with 70% ethanol. Place bubble wrap in the bottom of the box and cover with paper.
Separate mice into groups of three, placing in individual holding cups.
Place first mouse onto the center of the wire mesh and slowly invert 10-20 cm above the soft surface in the bottom of the Plexiglass box. Measure the latency to fall using a stopwatch. Terminate testing after 60 sec have elapsed if the mouse does not fall.
Return mouse to holding cup, wipe mesh with 70% ethanol and replace paper over the bubble wrap. Conduct the test on the remaining two mice in the group before returning to the first. Repeat until each mouse has been tested a total of three times.
Return the group to a clean cage and continue with remaining groups. If there are fewer than three mice in any group, ensure that a 5 min inter-trial recovery interval is allowed.
Return all mice to the home cage, dispose of paper and clean testing equipment with 70% ethanol.
Anaesthetize animals at P30-35 by intraperitoneal injection of ketamine (100 mg/kg) and xylazine (10 mg/kg). Assess depth of anesthesia by pinching the footpad and proceed only in the absence of a response, in line with IACUC guidelines. Perfuse with 4% PFA/PBS48.
To remove the eyes, clean the skin at the incision site with 70% ethanol, before cutting through the skin at the base of the skull. Peel back the skin to expose the skull and clean with 70% ethanol. Using clean instruments, carefully cut through the skull to reach the eye sockets and expose the eyes. Sever the optic nerve, gently remove the eye from the socket and rinse in PBS.
Fix eyes in 4% PFA/PBS for 10 min at room temperature.
Place eyes in a small petri dish containing PBS. Make a small incision in the center of the cornea using stitch removal scissors. Remove the cornea by cutting the edge between the cornea and sclera. Use forceps to remove the iris. Finally, peel off the RPE layers, leaving the retina intact and the lens in place.
Note: The 4% PFA/PBS fixation will cause RPE detachment allowing for the RPE to be easily peeled from the outside of the retina.
Re-fix in 4% PFA/30% sucrose/PBS for 15 min at room temperature before returning the eye to a petri dish containing PBS. Pull out the lens using forceps, bringing the vitreous body along with it.
Cryoprotect in 30% sucrose overnight at 4 °C.
Coat eyes in Optimum Cutting Temperature compound (OCT) and transfer to an embedding mold containing fresh OCT. Orient the eye such that the sagittal plane is parallel with the cutting face, using as few movements as possible and avoiding the introduction of air bubbles. Flash freeze by immersing the mold in a dry ice/ethanol bath. Store blocks at -80 °C.
Using a cryostat, cut 20 µm sagittal sections and collect on precleaned slides.
Stain sections with hematoxylin and eosin according to standard protocols49. Image using a light microscope equipped for bright field microscopy.
Euthanize animals at P30-35 using isoflurane overdose (30% in propylene glycol) via open drop inhalation. Assess euthanasia by the cessation of breathing, heartbeat and lack of response to pinching the footpad. Confirm death by cervical dislocation and organ removal, in line with IACUC guidelines.
To remove the brain, clean the skin at the incision site with 70% ethanol, before cutting through the skin at the base of the skull. Peel back the skin to expose the skull and clean with 70% ethanol. Using clean instruments, carefully cut through the skull and remove to expose the brain. Remove the membranes, and gently remove the brain from the skull cavity. Rinse in PBS.
Coat the brain in OCT and transfer to an embedding mold containing fresh OCT. Orient the brain such that the sagittal plane is parallel with the cutting face, using as few movements as possible and avoiding the introduction of air bubbles. Flash freeze by immersing the mold in a dry ice/ethanol bath and store at -80 °C.
Using a cryostat, cut 10 µm sagittal sections and collect on precleaned slides.
Prepare 0.05 M Tris buffer, pH 7.6 by dissolving 5.72 g Tris-HCl and 1.66 g Tris base in 1L-deionized water. Store at 4 °C for up to six weeks.
Prepare NADH solution (2.25 mM NADH in 0.05 M Tris buffer). Aliquot and store at -20 °C.
Prepare NBT solution (2.5 mM Nitro-Blue Tetrazolium in 0.05 M Tris buffer). Store at 4 °C for up to six weeks in a glass bottle.
Combine equal volumes of NADH solution and NBT solution to make staining solution and add 1 ml to each slide. Incubate at 37 °C in a humidified chamber for 30 min.
Aspirate staining solution and wash three times with dH2O.
Wash three times each in ascending and descending concentrations of acetone/dH20 (30%, 60%, 90%, 60%, 30%) for 2 min at room temperature.
Rinse three times in dH2O and mount sections with aqueous medium. Image using a light microscope equipped for bright field microscopy. Blue staining corresponds to enzymatic activity.
Prepare 0.2 M sodium monobasic phosphate/dH2O and 0.2 M sodium dibasic phosphate/dH2O.
Make 0.2 M phosphate buffer, pH 7.6 by combining 3 parts monobasic phosphate with 20 parts sodium dibasic phosphate.
Prepare staining solution (0.1 M sodium succinate, 1.25 M NBT in 0.2 M phosphate buffer) and add 1 ml to each slide. Incubate at 37 °C in a humidified chamber for 60 min.
Aspirate staining solution and wash three times in dH2O.
Wash three times each in ascending and descending concentrations of acetone/dH2O (30%, 60%, 90%, 60%, 30%) for 2 min at room temperature.
This section describes examples of the results obtainable using these methods. In the hind-limb test, the number of pull attempts made and the latency to fall are summed over two consecutive tests for each day. This test can be used to compare genetically different groups to distinguish mice with reduced neuromuscular strength. HtrA2tm1jhoh (HTRA2 KO) mice in Figure 1A-B demonstrate no change in the number of pulls and latency to fall from P4-6 followed by a reduction in neuromuscular strength at P7-10, as compared to littermates (HTRA2 WT). The expected increase in strength with age is also observable in the wild-type animals. For the weanling observation test, total activity can be evaluated by summing all events (lines traversed, rearing and grooming events), averaged across the three consecutive days of testing. Alternatively, each specific type of event can be individually analyzed. Here representative results are presented, demonstrating reduced activity of HTRA2 KO mice compared to WT littermates. These data demonstrate that total activity can be quantitatively measured and compared across groups (Figure 1C). It is also possible to quantify differences in grip strength by calculating a daily average from the three repeats performed on each testing day. When HTRA2 KO mice are compared to littermate controls, reductions in strength manifest in a shorter latency to fall (Figure 1D).
H&E staining of the retina generates images in which retinal structure can be examined and compared (Figure 2). The layers of cell types that compose the retina are clearly delineated allowing comparison between genetic groups. Here, HtrA2tm1jhoh';HtrA3tm1jhoh mice show no changes in retinal structure, compared to HtrA2WT;HtrA3tm1jhoh mice at P35. Finally, histochemical staining for electron transport chain complex activity can be used to identify changes in respiratory enzyme functions, with increases in staining intensity reflecting increase enzyme activity, such as the reduction observed in the cerebellum and striatum of HtrA2flox/flox;Tg(Nes-cre)1Kln/J (NesKO) brain when compared to WT littermates (Figure 3).
Figure 1: Behavioral phenotyping of genetically engineered mice. HtrA2tm1jhoh (HTRA2 KO) mice were subjected to the described behavioral testing and compared to wildtype (HTRA2 WT) littermate controls. (A-B) The hind limb suspension test was performed on WT and KO neonates from P4-P10 and demonstrates a reduction in neuromuscular strength in KO animals (WT: n = 28, KO: n = 19). (A) KO pups initially make a similar number of pull attempts as WT siblings, but show a decrease in attempts made at later time points. (B) KO neonates' latency to fall is also initially normal but decreases from P7 onwards. (C) The weanling observation total activity score was reduced in KO animals compared to WT littermates, consistent with reduced activity of these mice (WT: n = 19, KO: n = 12). (D) KO animals also had a reduced latency to fall during the mesh inversion test, demonstrating reduced grip strength of HTRA2 KO mice (WT: n = 17, KO: n = 9). Data represents Mean ± SEM (# 0.1 <p <0.05, *p <0.05, **p <0.001, ***p <0.001 by independent t-tests). This figure has been modified from Patterson et al.34 Please click here to view a larger version of this figure.
Figure 2: H&E staining of retina. Staining retinal sections with hematoxylin and eosin allows examination of structure, delineating the retinal layers. Here sections from P35 HtrA2WT;HtrA3tm1jhoh (A) and HtrA2tm1jhoh;HtrA3tm1jhoh (B) display normal retinal structure. RPE: retinal pigment epithelium, OS: outer segments, IS:inner segments, ONL: outer nuclear layer, OPL: outer plexiform layer, INL: inner nuclear layer, IPL: inner plexiform layer, GCL; ganglion cell layer. Scale bar = 1 mm. Please click here to view a larger version of this figure.
Figure 3: Respiratory complex activity. Histochemical staining for NADH diaphorase (complex I) and succinate dehydrogenase (complex II) activity was performed on transverse brain sections of P25 HtrA2flox/flox;Tg(Nes-cre)1Kln/J (NesKO) and HtrA2+/flox;Tg(Nes-cre)1Kln/J (NesWT) littermates. NADH diaphorase enzyme activity is decreased in cerebellum (Crbl, B) and striatum (D) of NesKO brain compared to NesWT controls (A, C), but not in cerebral cortex (E-F). SDH activity is similarly reduced in cerebellum and striatum of NesKO mice (H, J) compared to NesWT controls (G, I) but there is no change in cerebral cortex (K-L). Scale bar = 200 µm. This figure has been modified from Patterson et al.34 Please click here to view a larger version of this figure.
Robust treatments are needed to limit the impact of debilitating conditions related to human aging, but they remain elusive for many conditions. To identify potential therapeutic targets and develop strategies for intervention, the causes and pathology of diseases associated with aging must first be understood. Not all genetically modified mice immediately present with clear phenotypes that are related to the disease of interest, even if those genes have previously been linked to the condition in human studies. Therefore more systematic investigations are required and genetically modified mice should be subjected to appropriate experiments to identify phenotypes that may be related to the associated diseases in humans.
Myriad behavioral tests exist to assay neurological function in mice50,51. The behavioral tests outlined herein are suited to the hypothesized phenotypes based on human findings and the actual phenotypes observed in mice. In addition, these tests are simple to perform in laboratories with little or even no experience in conducting behavioral experiments. There is no requirement for sophisticated equipment but the results are meaningful and informative when properly collected, making them useful as a first step in examining the phenotype of genetically modified mice that are expected to have neuromuscular defects. The resultant data can then be used to direct the focus of further testing, potentially with more sophisticated testing apparatus.
Similarly, many techniques exist for assessing respiratory function in tissues52. Measuring the rate of ATP synthesis, oxygen consumption or changes in mitochondrial membrane potential are important readouts of mitochondrial function but they can be complicated and time consuming to perform, requiring specific reagents and equipment52-55. In addition, they are often limited to the level of the mitochondrion or cell, relying on cultured cells, lysates or isolated mitochondria. Histochemical staining is a useful first step in identifying mitochondrial dysfunction without a large investment of time or resources. Moreover, the ability to stain tissue sections lends itself to quickly identifying regional differences that might not be as apparent with other techniques, for example the area-specific loss of enzymatic staining observed in HtrA2-deficient brain. While the technique is limited in its ability to measure fold changes in respiratory capacity, it maps out defined brain regions that can then be pursued using more complex methods.
While the techniques presented within this phenotyping schedule are simple to conduct, performing certain steps properly is critical to the success of the experiments. For the behavioral phenotyping, great care must be taken to ensure tests are performed at the same time and in the same location each day to limit variation introduced by the novelty of location or by the difference in the circadian rhythm. The equipment should be organized in the same way each day for similar reasons. The room organization is especially critical for the activity test, where small changes in novelty can greatly influence the activity of the mouse. When measuring grip strength, investigators should ascertain that mice have a proper hold of the mesh before inversion. Similarly, care must be taken to properly suspend the pup before releasing in the hind limb test or the test results will be skewed towards a shorter latency to fall.
For tissue phenotyping, obtaining good sections is vital. Informative staining of the retina depends upon effective fixation before dissection or artifacts will arise during sectioning and dissection and distort the morphology. Histochemical staining requires brains to be frozen quickly after dissection to preserve enzyme function, but the incubation time is similarly critical; small fluctuations in temperature or in the length of time can cause differences in the staining intensity. Where possible, samples should be processed at the same time to allow comparison between groups.
Here a testing regimen used to study genetically modified mice expected to display subtle phenotypes associated with age-related diseases, namely AMD and PD, is described. This phenotyping schedule can be used to identify deficits in neuromuscular strength and activity, as well as identify structural and functional defects within the tissues of interest. This schedule provides a systematic analysis of early phenotypes in young, genetically altered mice. In the future, this method can be applied to any genetically modified mice expected to present with an AMD or PD related phenotype, as well as those of unknown etiology that appear overtly normal. The simplicity of the tests allows testing without significant investment but can be used to inform further studies. By combining behavioral testing with cellular phenotyping, it is possible to limit the number of mice used and directly compare animal performance with the physiological condition. This phenotyping schedule provides an initial analysis that can be used to direct the focus of subsequent studies.
Funding for this research came from Rosebay Medical Foundation and a Yale Medical School Dean's Research Fund (JH). We thank Dr. Claire Koenig for help with behavioral experiments. Genetically engineered mouse lines were generated at Ozgene (Perth, Australia).
Plexiglass box constructed in workshop 12" by 12"
1/4 inch wire mesh Home Depot 204331884 (online) / 554219 (in store) 12" by 12"
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Sicilian Ghost Story's otherworldly nature and the powerful subject matter is deeply impactful.
Out this week is Sicilian Ghost Story directed by Fabio Grassadonia and Antonio Piazza. This film is based on true events involving the kidnapping of a young boy from a village due to his father's betrayal of an Italian mafia family.
The film centres around two characters, Giuseppe (Gaetano Fernandez) the young boy who is kidnapped and Luna (Julia Jedlikowska) who is a fictional character injected into the story to give, what the directors felt Giuseppe deserved, love. These two are best friends with a clear love of each other and when Giuseppe is kidnapped Luna is the only one to try and figure out what happened. The film follows Luna as she tries to figure out what to do all the while giving the audience a parallel storyline with Giuseppe and his captors.
There is a theme of rebellion that streaks through the film and its characters. From Giuseppe and his captors to Luna and her overbearing mother, the fire of rebellion is strong in the characters. This is needed for both characters as it is what helps them both survive in their respective prisons.
The biggest positive I can offer this film is in the shape of its two young leads. They are astounding. Both give deeply moving performances. The film demands so much from them but they rise to the challenge and leave a lasting impression. As well as the Giuseppe and Luna the intimate cast also has some interesting performances including a character Luna meets in the second act. A young boy who for a brief moment lets her forget that someone she deeply cares about is missing. it's this brief respite that allows for the film to balance between its graceful yet brutal nature.
There is also the impressive cinematography that continues the brutal yet artistic grace of the film. The scenery, in particular, is gorgeous and has an otherworldly element to it. Coupling it with the dual narrative of what Luna imagines is happening to Giuseppe and what is actually happening to Giuseppe adds a tense element to the film.
The symbolism and surreal nature of Sicilian Ghost Story are commendable but at times they detract from the overall quality of the film because you may not understand what's happening. Whether something is real or not is not always explained well and it's a shame because it took me out of the film several times to try and figure it out.
Sicilian Ghost Story is a love letter to a young unloved boy who fell victim to a horrifying time in Italian history by two brilliant directors. It's otherworldly nature and the powerful subject matter is deeply impactful.


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With an ocean of information out there about risk, it's likely that you're already feeling the pull of the tide. Nowadays internal audit teams are frequently being requested to take a more consultative approach and assume a stronger focus on risk management. Sound familiar? This is because internal audit departments are in a unique position to help business leaders comprehend and navigate risk. And as organizations continue to navigate increasingly complex business environments, audit's role is evolving and risk acumen is vital.
Although risk knowledge can be overwhelming, let's get one thing straight–not all risk is bad. A complete absence of business risk virtually guarantees limited growth. Taking risks within your organization's risk tolerance and risk appetite can help your organization grow and achieve your goals. The key is to understand your organization's risk appetite before you can audit it.


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