the four experiments presented here can be performed independently or as a four - part series .
experiment 1 introduces students to the concept of model prebiotic chemical reactions , experiment 2 teaches the students how to isolate dna from a living organism , experiment 3 teaches the students how to release purine bases from dna by the acid - catalyzed depurination reaction , and experiment 4 teaches the students about tlc .
adenine , produced chemically in experiment 1 and derived from a biological source in experiment 3 , is easily identified by the use of tlc and authentic standards .
thus , the teacher may choose to conduct experiment 4 first as a means to introduce students to the tlc technique , so that students are able to use tlc in experiment 1 and experiment 3 .
however , if time is limited , the teacher may opt to demonstrate the tlc technique rather than having students perform the tlc procedure separately .
if this option is selected , the correct setup and purpose of the various parts of a tlc chamber are described below in experiment 4 and in the supporting information .
students use plausible prebiotic molecules to synthesize a nitrogenous base ( adenine ) that is found in the dna of all living organisms . hydrogen cyanide ( or hcn )
has long been considered a precursor to nitrogenous bases because hcn is a simple molecule that could have been produced in the atmosphere of the early earth.(2 ) one molecule of hcn will react with one molecule of water to produce formamide , also a likely prebiotic molecule ( scheme 1 ) .
moreover , formamide can further react with water to produce ammonium formate , a salt ( scheme 1 ) .
the accepted relevance of hcn , formamide , and ammonium formate to prebiotic chemistry is also supported by the presence of water , hcn , and formamide in interstellar space .
the molecule diaminomaleonitrile , or damn ( scheme 2 ) , is also known as the hcn tetramer because it is made up of four hydrogen atoms , four carbons , and four nitrogens .
damn is considered a potentially important molecule of prebiotic chemistry because it is an intermediate in the reaction pathway from concentrated hcn to adenine.(22 ) inspiration for this experiment came from the discoveries that small quantities of adenine are formed in heated solutions of formamide and that adenine is generated in good yield when damn is heated in molten ammonium formate ( scheme 3).(4 ) these reactions are considered relevant to understanding prebiotic chemistry and astrochemistry because formamide , ammonium formate , and damn are all products of hcn chemistry.(4 ) to make this chemistry accessible to the high school classroom , a timesaving , simplified procedure for the synthesis of adenine in a model prebiotic reaction has been developed .
students perform a reaction in which ammonium formate is dissolved in formamide ; damn is then added , and the reaction is heated to accomplish the formation of adenine .
the students compare , by tlc , the nitrogenous base synthesized in this reaction to authentic standards of adenine , thymine , and cytosine .
guanine is not used as a standard in these experiments because it has a low solubility in water .
protocols for extracting dna from different plants , fruits , and vegetables are available online,(23 ) but the goal of these various protocols is usually for students to visualize chromosomal dna with the naked eye . a protocol is included for dna extraction that allows students to use the extracted dna in the hydrolysis reaction of an additional laboratory experiment in this series .
strawberries are mashed in a dna extraction buffer made of sodium chloride , clear liquid soap , and water .
frozen strawberries may also be thawed and used if fresh ones are not available ; overripe strawberries are also acceptable .
the strawberry mixture is then filtered using filter paper or cheesecloth to remove the solids before chilled ethanol is poured down the side of the container with the filtered strawberry mixture .
ninety - five percent ethanol ( denatured ) is preferred for this step , although 70% ethanol will also suffice .
concentrations of ethanol lower than 70% should not be used . the dna obtained from this step
this experiment consists of two parts , each designed to take one 4560 min lab period . in the first part ,
students dissolve their extracted dna from experiment 2 in water and heat this solution in a water bath to dissociate the dna from histones and other bound proteins .
the heated dna solution is cooled , and 3 m hydrochloric acid is added ; the solution is again heated in a water bath . both heating steps are essential for liberated adenine to be detected using the tlc analysis protocol provided in experiment 4 .
the first , acid - free heating step must be performed separately from the acidic heating step .
adenine and guanine , the purine nucleobases of dna ( and rna ) , become protonated in acidic solution , which facilitates the hydrolysis of the bond between these nucleobases and the sugar  phosphate backbone of dna , a process known as depurination .
part two of this experiment will need to be completed within one or two days of part one . in part two
, tlc is used to analyze the partially hydrolyzed dna . because guanine has very low solubility in water
the adenine will migrate with the solvent up the plate and can be compared to authentic standards .
this experiment is designed for teachers to implement in classrooms where they feel the students would benefit from extra practice with the tlc technique before moving on to experiment 1 .
note that if experiment 4 is not performed , the time it takes to complete experiment 1 will likely be greater as students may need to gain proficiency with tlc . if the tlc demonstration option is chosen , in place of completing experiment 4 , instructions for preparing and running a tlc plate are provided in experiment 1 as a reminder to students .
however , if time permits , students may practice the tlc technique themselves by doing this experiment before beginning experiment 1 .
students are instructed in the basics of preparing tlc chambers and plates and calculation of retention factors ( rf ) .
students will run authentic standard solutions of adenine , thymine , and cytosine on their first tlc plate , and various mixtures of these compounds , prepared by the instructor , on their second tlc plate .
they will then identify the unknowns from the mixtures based on their results with authentic standards . a piece of filter paper is cut to dimensions that will fit into a plastic cup or beaker ( the tlc chamber ) , reaching to the top , while covering one - half or less of the chamber wall surface area .
solvent , in this case , water , is placed in the chamber , taking care to keep the level below that of the origin line where the analyte spots are placed on the tlc plate .
the chamber is covered with a watch glass or with plastic wrap and a rubber band .
the origin line is marked with a pencil and ruler on the tlc plate , and marks are made where each analyte will be placed .
spots are carefully made on the plate using a glass capillary tube or flat toothpick . when dry , the plate is placed into the chamber and the water
is allowed to travel most of the way up the plate , which is then removed from the chamber with forceps .
the plate is dried and then visualized with shortwave uv light ( 254 nm ) .
uv lamps , used for visualization of analytes on fluorescent tlc plates , should always face downward , and students should take care not to look directly into them .
caution should be exercised in handling hot containers after the model prebiotic reaction in experiment 1 and during the dna hydrolysis in experiment 3 .
glass capillary tubes used for tlc analyte spotting may be sharp , and students should not dispose of these tubes in the regular trash where cleaning staff may encounter them .
( glass capillary tubes may be placed into a secondary container , such as an empty plastic water bottle , before disposal . ) because water is the eluting solvent , there are no hazards associated with the tlc developing chambers or with residual solvents on the tlc plates . similarly , the dna extraction buffer from experiment 2 contains safe ingredients and may be disposed of by pouring down the drain .
adenine , cytosine , and thymine are listed as irritants in crystalline form , but will only be used in very dilute solutions in this experiment series . if these solutions touch the skin , washing with soap and water is sufficient to remove nucleobases and prevent irritation .
formamide , ammonium formate , and damn are toxic by ingestion and inhalation , and therefore care should be taken to prevent contact of these chemicals with the skin or eyes .
hydrochloric acid , 3 m , is a strong acid and should be treated with extreme caution .
experiments have been developed that allow prebiotic chemistry to be brought into the high school classroom . by participating in the series of low - cost laboratory experiments , using tlc as a means to identify compounds in biological and chemical samples ,
students gain a deep appreciation for the connections that exist between contemporary biochemistry ( e.g. , the molecular structure of dna ) , molecules that are widespread in interstellar space , and how these components may have provided the chemical inventory that gave rise to biopolymers on the early earth .
 many high school laboratory experiments demonstrate concepts related to biological evolution , but few exist that allow students to investigate life s chemical origins . 
 this series of laboratory experiments has been developed to allow students to explore and appreciate the deep connection that exists between prebiotic chemistry , chemical evolution , and contemporary biological systems . in the first experiment of the series , 
 students synthesize adenine , one of the purine nucleobases of dna and rna , from plausibly prebiotic precursor molecules . 
 students compare their product to authentic standards using thin - layer chromatography . 
 the second and third experiments of the series allow students to extract dna from a familiar organism , the strawberry , and hydrolyze it , releasing adenine , which they can then compare to the previously chemically - synthesized adenine . a fourth , optional experiment 
 is included where the technique of thin - layer chromatography is introduced and chromatographic skills are developed for use in the other three experiments that comprise this series . 
 concepts relating to organic and analytical chemistry , as well as biochemistry and dna structure , are incorporated throughout , allowing this series of laboratory experiments to be easily inserted into existing laboratory courses and to reinforce concepts already included in any high school chemistry or biology curriculum . 
Procedure and Results
Hazards
Conclusion