中国人兽共患病学报
中國人獸共患病學報
중국인수공환병학보
CHINESE JOURNAL OF ZOONOSES
2010年
3期
235-238,242
,共5页
卡氏肺孢子菌%p55-v0及p55-v3%基因克隆%序列分析
卡氏肺孢子菌%p55-v0及p55-v3%基因剋隆%序列分析
잡씨폐포자균%p55-v0급p55-v3%기인극륭%서렬분석
Pneumocystis carinii%p55-v0 and p55-v3%gene cloning%sequence analysis
目的 扩增卡氏肺孢子菌p55-v0及p55-v3基因的CDS区,并对其进行序列分析比较.方法 从感染卡氏肺孢子菌的鼠肺组织中提取总RNA,以其为模板采用RT-PCR法分别扩增p55-v0和p55-v3编码基因,然后将相应PCR产物与T载体连接,转化感受态DH5α,在含氨苄青霉素的LB培养基上筛选重组T载体,PCR扩增,酶切,测序并进行序列比较.结果 p55-v0的CDS区基因长度为 1 245bp,编码414个氨基酸,比较发现其与文献报道的同源性分别为99.8%和100%.p55-v3 CDS区基因长度为1 053bp,编码350个氨基酸,与GenBank同源性分别为99.9%和100%.然而p55-v0与p55-v3基因之间的同源性仅20.9%.结论 本研究成功地克隆了p55-v0和p55-v3基因CDS区,分析发现p55-v0和p55-v3与国外报道的存在很高的同源性,但它们两者之间在基因组成及氨基酸组成上存在很大的差异,这为进一步比较它们的免疫功能从而阐明p55-v3的作用机制提供了基础.
目的 擴增卡氏肺孢子菌p55-v0及p55-v3基因的CDS區,併對其進行序列分析比較.方法 從感染卡氏肺孢子菌的鼠肺組織中提取總RNA,以其為模闆採用RT-PCR法分彆擴增p55-v0和p55-v3編碼基因,然後將相應PCR產物與T載體連接,轉化感受態DH5α,在含氨芐青黴素的LB培養基上篩選重組T載體,PCR擴增,酶切,測序併進行序列比較.結果 p55-v0的CDS區基因長度為 1 245bp,編碼414箇氨基痠,比較髮現其與文獻報道的同源性分彆為99.8%和100%.p55-v3 CDS區基因長度為1 053bp,編碼350箇氨基痠,與GenBank同源性分彆為99.9%和100%.然而p55-v0與p55-v3基因之間的同源性僅20.9%.結論 本研究成功地剋隆瞭p55-v0和p55-v3基因CDS區,分析髮現p55-v0和p55-v3與國外報道的存在很高的同源性,但它們兩者之間在基因組成及氨基痠組成上存在很大的差異,這為進一步比較它們的免疫功能從而闡明p55-v3的作用機製提供瞭基礎.
목적 확증잡씨폐포자균p55-v0급p55-v3기인적CDS구,병대기진행서렬분석비교.방법 종감염잡씨폐포자균적서폐조직중제취총RNA,이기위모판채용RT-PCR법분별확증p55-v0화p55-v3편마기인,연후장상응PCR산물여T재체련접,전화감수태DH5α,재함안변청매소적LB배양기상사선중조T재체,PCR확증,매절,측서병진행서렬비교.결과 p55-v0적CDS구기인장도위 1 245bp,편마414개안기산,비교발현기여문헌보도적동원성분별위99.8%화100%.p55-v3 CDS구기인장도위1 053bp,편마350개안기산,여GenBank동원성분별위99.9%화100%.연이p55-v0여p55-v3기인지간적동원성부20.9%.결론 본연구성공지극륭료p55-v0화p55-v3기인CDS구,분석발현p55-v0화p55-v3여국외보도적존재흔고적동원성,단타문량자지간재기인조성급안기산조성상존재흔대적차이,저위진일보비교타문적면역공능종이천명p55-v3적작용궤제제공료기출.
This study sought to amplify the CDS region of p55-v0 and p55-v3 gene in Pneumocystis carinii (PC) and to analyze their sequences.The total RNA extracted from PC- infected lungs of rats were used as the template to amplify these genes by RT-PCR.The amplified products were connected to T-vector and transformed into E.coli DH5α.The recombinant T-vectors were selected in LB culture medium containing ampicillin.Then the positive clones of recombinants were identified by PCR and digested by restrictive endonuclease.After that the recombinant were sequenced and analyzed.It was found that the CDS region of p55-v0 gene contained 1245 bps,encoding 414 amino acids.The homology between this sequence and the one reported previously in nucleotide and amino acid were 99.8% and 100% respectively.Meanwhile,the CDS region of p55-v3 contained 1053 bps,encoding 350 amino acids.As compared with GenBank,the homology in nucleotide and amino acid was 99.9% and 100% respectively.However,the similarity in nucleotide between p55-v3 and p55-v0 was just 20.9%.In this study,the CDS regions of p55-v0 and p55-v3 gene were cloned successfully,and the high homology was found between p55-v0,p55-v3 and the one reported previously by sequence analysis,p55-v3 was different from p55-v0 in nucleotides and amino acids.This result would provide a basis for comparison of immunologic functions so as to explore the mechanism of action of the p55-v3 gene.
