论文部分内容阅读
克隆中国对虾(Fenneropenaeus chinensis)胰蛋白酶基因,参考多种生物的胰蛋白酶基因序列设计出1对简并PCR引物。利用PCR技术从中国对虾基因组DNA中扩增出1条DNA带,经PCR产物回收、纯化,连接到载体质粒,转化细菌,蓝百斑筛选等操作获得了1个阳性克隆。序列分析表明,该序列包含1个内含子和2个不完整的外显子,与已报道的凡纳滨对虾(Litopenaeus vannamei)胰蛋白酶基因Ⅲ有较高的相似度(identity=79.2%)。综合多种分析,可以推定该序列为对虾的胰蛋白酶基因序列。系统进化分析表明该基因属于胰蛋白酶第三家族。
One pair of degenerate PCR primers was designed by cloning the trypsin gene of Fenneropenaeus chinensis and referring to the trypsin gene sequences of various organisms. One DNA band was amplified from genomic DNA of Penaeus chinensis by PCR, and one positive clone was obtained after PCR products were recovered, purified, ligated into vector plasmid, transformed into bacteria and screened by blue dot blot. Sequence analysis showed that the sequence contained one intron and two incomplete exons, which had a higher identity (identity = 79.2%) to that reported in Litopenaeus vannamei trypsin gene Ⅲ . Based on a variety of analyzes, it can be inferred that the sequence is the trypsin gene sequence of shrimp. Phylogenetic analysis showed that this gene belonged to the third family of trypsin.