中国动物传染病学报
中國動物傳染病學報
중국동물전염병학보
CHINESE JOURNAL OF VETERINARY PARASITOLOGY
2014年
5期
41-48
,共8页
王晓娟%米荣升%周鹏%黄燕%王向佩%杨晓娇%石凯%刘宇轩%雷晓思%陈兆国%赵权
王曉娟%米榮升%週鵬%黃燕%王嚮珮%楊曉嬌%石凱%劉宇軒%雷曉思%陳兆國%趙權
왕효연%미영승%주붕%황연%왕향패%양효교%석개%류우헌%뢰효사%진조국%조권
隐孢子虫%Cryptopain-1基因%克隆%序列分析
隱孢子蟲%Cryptopain-1基因%剋隆%序列分析
은포자충%Cryptopain-1기인%극륭%서렬분석
Cryptosporidium%cryptopain-1 gene%cloning%sequence analysis
为克隆不同种隐孢子虫（Cryptosporidium spp.）的半胱氨酸蛋白酶Cryptopain-1基因，分析其核苷酸与编码氨基酸序列的变异情况，根据GenBank上公布的微小隐孢子虫（C. parvum）Cryptopain-1基因序列设计合成引物，用PCR技术从不同种隐孢子虫基因组DNA中扩增Cryptopain-1基因，并将其克隆至pZeroBack/blunt载体，阳性克隆经PCR鉴定正确后测序，与微小隐孢子虫Cryptopain-1基因进行序列同源性比对，并进行系统进化分析。结果显示，本研究成功从泰泽隐孢子虫（C. tyzzeri）、火鸡隐孢子虫（C. meleagridis）、兔隐孢子虫（C. cuniculus）基因组DNA中扩增出Cryptopain-1基因。与微小隐孢子虫Cryptopain-1基因比较，泰泽隐孢子虫、火鸡隐孢子虫、兔隐孢子虫Cryptopain-1基因核苷酸序列相似性分别为98.67%、94.53%、98.34%，演绎的氨基酸序列相似性分别为99.00%、96.51%、99.00%。研究结果为利用Cryptopain-1基因进行隐孢子虫的代谢、致病机理等研究奠定了基础。
為剋隆不同種隱孢子蟲（Cryptosporidium spp.）的半胱氨痠蛋白酶Cryptopain-1基因，分析其覈苷痠與編碼氨基痠序列的變異情況，根據GenBank上公佈的微小隱孢子蟲（C. parvum）Cryptopain-1基因序列設計閤成引物，用PCR技術從不同種隱孢子蟲基因組DNA中擴增Cryptopain-1基因，併將其剋隆至pZeroBack/blunt載體，暘性剋隆經PCR鑒定正確後測序，與微小隱孢子蟲Cryptopain-1基因進行序列同源性比對，併進行繫統進化分析。結果顯示，本研究成功從泰澤隱孢子蟲（C. tyzzeri）、火鷄隱孢子蟲（C. meleagridis）、兔隱孢子蟲（C. cuniculus）基因組DNA中擴增齣Cryptopain-1基因。與微小隱孢子蟲Cryptopain-1基因比較，泰澤隱孢子蟲、火鷄隱孢子蟲、兔隱孢子蟲Cryptopain-1基因覈苷痠序列相似性分彆為98.67%、94.53%、98.34%，縯繹的氨基痠序列相似性分彆為99.00%、96.51%、99.00%。研究結果為利用Cryptopain-1基因進行隱孢子蟲的代謝、緻病機理等研究奠定瞭基礎。
위극륭불동충은포자충（Cryptosporidium spp.）적반광안산단백매Cryptopain-1기인，분석기핵감산여편마안기산서렬적변이정황，근거GenBank상공포적미소은포자충（C. parvum）Cryptopain-1기인서렬설계합성인물，용PCR기술종불동충은포자충기인조DNA중확증Cryptopain-1기인，병장기극륭지pZeroBack/blunt재체，양성극륭경PCR감정정학후측서，여미소은포자충Cryptopain-1기인진행서렬동원성비대，병진행계통진화분석。결과현시，본연구성공종태택은포자충（C. tyzzeri）、화계은포자충（C. meleagridis）、토은포자충（C. cuniculus）기인조DNA중확증출Cryptopain-1기인。여미소은포자충Cryptopain-1기인비교，태택은포자충、화계은포자충、토은포자충Cryptopain-1기인핵감산서렬상사성분별위98.67%、94.53%、98.34%，연역적안기산서렬상사성분별위99.00%、96.51%、99.00%。연구결과위이용Cryptopain-1기인진행은포자충적대사、치병궤리등연구전정료기출。
In order to clone Cryptopain-1 gene of different Cryptosporidium species for analysis of genetic variation, a pair of specific primers were designed based on the sequence of C. parvum Cryptopain-1 gene published on GenBank. The Cryptopain-1 genes of different Cryptosporidium species were amplified from their genomes using PCR technique and then cloned into pZeroBack/blunt vectors. The cloned genes from the positive recombinant plasmids were sequenced and phylogenetically analyzed. Cryptopain-1 gene of C.parvum shared nucleotide identities at 98.67%, 94.53%and 98.34%with C.tyzzeri,C.meleagridis and C.cuniculus. The information obtained from the present study will be useful for research on metabolism, pathogenecity of Cryptosporieium.
