Pagrosomus major antibacterial peptide gene, and recombinant yeast expression vector and its preparing method
A yeast expression vector, recombinant yeast technology, applied in the fields of botanical equipment and methods, biochemical equipment and methods, genetic engineering, etc., can solve problems such as unreported
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[0030] Taking the cloning of the red sea bream antimicrobial peptide hepcidin gene and the construction of the expression vector as an example, the embodiments of the present invention will be described in detail in conjunction with the accompanying drawings.
[0031] 1. Construction of cDNA library: The cDNA library was constructed at the Yellow Sea Fisheries Research Institute from April to July 2002. Total RNA was extracted from red sea bream spleen with TRIzol kit, and Oligotex TM The spin column kit was used to separate Poly-(A) RNA, and the pBluescript II XR cDNA library kit was used to construct a cDNA library containing 1.5×10 6 clone.
[0032]2. EST sequence analysis and red sea bream antimicrobial peptide gene sequence: Plasmid DNA was prepared by a rapid micropreparation method, and the plasmid preparation was completed on a 96-well plate; the sequence of the cDNA fragment was determined using a T3 sequencing kit, and T3 and T7 primers were used to obtain the seque...
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