Screening And Identification Of Antimicrobial Peptide Gene In Pengze Crucian Carp

The outbreak of infectious diseases have caused huge losses in fish culture industry. As the law against antibiotics residue has becoming more and more stringent, the antibiotics application in the fish culture industry is greatly restricted. The production of antimicrobial peptides(AMPs) has been identified as a major defense mechanism against infections. So AMPs are regarded as well substitution of antibiotics. Pengze Crucian Carp is the first cultural breed variety which is artificially and directly selected from wild crucian. Pengze Crucian Carp has many good characteristics,for example,it is easy to culture,it grows fast,it has high nutritive value and it has strong antireversion force and so on, which make it one of the most important fresh water commercial fishes. We propose to find new AMPs genes from Pengze Crucian Carp and to establishe the foundation for the exploitation of new Biological Pesticide.High quality total RNA were isolated from healthy Pengze Crucian Carp (C-group) and Aeromonas hydrophila induced Pengze Crucian Carp (T-group). The differential expressed genes in liver of C-group and T-group were studied via DDRT-PCR technique. Then we utilized the PAGE incorporate with silver staining to displayed the DDRT-PCR products and obtained 31 differential displayed bands which are specially expressed by T-group .After re-amplification, PAGE identification and purification of the 31 differential displayed bands, we got 20 differential displayed cDNA fragments. We cloned the 20 differential displayed cDNA fragments into E.Coli DH5-αusing T-A cloning technology and entrusted to Shanghai Sangon Biological Engineering Technology&Service Co.Ltd for sequencing and got 11 differential displayed ESTs sequences. We submitted the 11 new ESTs sequences to the dbEST of GenBank and their accession numbers are GE342614, GE342615, GE342616, GE342617, GE653346, GE653347, GE653348, GE653349, GH159107, GH159108, GH159109.We utilized the BLASTn and BLASTx tools to align the 11 new ESTs with different sequence databases of GenBank and acquired many homologous sequences, which offered references for the study of function of the new ESTs and their corresponding genes.