家蝇幼虫脂肪体cDNA文库的构建及初步鉴定

Construction and preliminary identification of the cDNA library for Musca domestica larvae

To construct the cDNA library for Musca domestica larvae, the total RNA was extracted from the fat globes of M.domestica larvae 24 hours after immuno induction with mixed bacterial suspensions of E.coli and Staphylococcus aureus.It was retro transcribed to synthesize the single strand cDNA using SMART oligo, and then synthesized and amplified double strand cDNA by long distance PCR(LD PCR). The PCR products were ligated into λTripIE x 2 vector, and the unamplified library was not completed until the recombinants were packaged by packing extracts. The results showed that the primary titer of the constructed cDNA library was 1.34×10 6 pfu/ml, while that of the amplified library was 2.408×10 9 pfu/ml.The recombination rate was about 98.9% and the length of most cDNA inserted in the library was about 400-1500 bp.It concludes that the cDNA library of Musca domestica larvae has been constructed by using the SMART technique.