嗜水气单胞菌诱导的中华鳖SMART cDNA文库构建及相关基因的鉴定

Construction of a SMART cDNA Library of Trionyx sinensis Stimulated with Aeromonas hydrophila and Molecular Identification

To understand anti-infectious response to bacteria in the Chinese soft shell turtle（Trionyx sinensis）, a full length cDNA library was constructed for it by SMART technique experimentally infected with Aeromonas hydrophila. Firstly, to synthesize the double-strand cDNA using SMARTTMPCR cDNA systhesis kit. Then, the ds cDNA was separated into two parts based on the size distribution of amplified ds cDNA by agarose gel size fractionation. The part that shorter than 500 bp discards and the other one which longer than 500 bp was ligated to the pGEM-T vector. The ligation mixture was transformed into E. coli JM109 by electroporation. The cDNA library contained 4.3×10^5 independent clones with DNA inserts of 0.5～2 kb. The recombination rate was 92%. We sequenced 164 cDNA clones about 1 kb, 39 genes are found for the first time in reptiles, and classified in function.Immune-related genes 11; Catalytic genes 15; Cell signaling genes 6; Transport genes 3; Cell structure and growth genes 7; Unknown function genes 9. The cDNA library provides a valuable resource for functional genome research of the Trionyx sinensis,and it will form a fundamental base for further research in immunobiological and phylogenetic aspects of turtles as well as in host-microbe interactions.