All of the mRNA samples were diluted to 1000ng/μl. We took 12.5 μl subsamples of the diluted mRNA for reverse transcription to cDNA using an Advantage reverse transcription kit for PCR (Clontech, Clayton Vic, Australia). The reverse transcription and PCR activation procedures were carried using Eppindorf Mastercycler (Eppindorf, Mamburg, Germany) qPCR machines, following the recipe outlined by the reverse transcription kit handbook . A “buffers only” (i.e. no RNA and no Reverse Transcriptase) solution was included in the analyses as a negative control. We included the spider’s abdomens in the gene expression analysis for normalization against background expression of MaSp transcripts in other silk glands or abdominal tissue . All procedures were replicated three times for each individual spider.