本文關(guān)鍵詞： 南方水稻黑條矮縮病毒 S片段 S-基因 原核表達　出處：《華南農(nóng)業(yè)大學學報》2017年01期 　論文類型：期刊論文

【摘要】：【目的】探討南方水稻黑條矮縮病毒(Southern rice black-streaked dwarf virus,SRBSDV)安徽分離物(SRBSDVAn Hui-HN2)的遺傳特性,并獲得原核表達的P5-2蛋白�！痉椒ā縍T-PCR擴增SRBSDV S5片段,克隆、測序并進行序列分析。將S5-2基因插入原核表達載體,重組載體轉(zhuǎn)化大腸埃希菌并用IPTG誘導,Ni2+-NTA親和柱純化融合蛋白,SDS-PAGE分析P5-2蛋白的表達情況�！窘Y(jié)果】SRBSDV-An Hui-HN2 S5片段全長3 167 bp,包含S5-2基因全長612 bp,編碼204個氨基酸。序列比對結(jié)果顯示,SRBSDV-An Hui-HN2 S5片段與其他SRBSDV分離物S5片段的序列相似性極高,達99.0%~99.7%,而與斐濟病毒屬(Fijivirus)其他成員S5片段的序列相似性較低,僅為38.0%~71.3%;構(gòu)建的S5片段系統(tǒng)發(fā)育樹表明SRBSDV和RBSDV聚成1個分支,其中6個SRBSDV分離物聚成1個亞分支。原核表達獲得相對分子質(zhì)量約為47 000的重組蛋白,Western blot分析顯示,GST單抗能夠與重組融合蛋白發(fā)生特異性反應(yīng)�！窘Y(jié)論】SRBSDV各分離物之間親緣關(guān)系非常近,而與Fijivirus其他成員親緣關(guān)系較遠,原核表達獲得的融合蛋白為靶標蛋白。
[Abstract]:[objective] to study the genetic characteristics of Southern rice black-streaked dwarf virus (SRBSDV) isolated from Anhui Province, and obtain the P5-2 protein expressed in prokaryotic expression. [methods] SRBSDV S5 fragment was amplified by RT-PCR and cloned. S5-2 gene was inserted into prokaryotic expression vector. The recombinant vector was transformed into Escherichia coli and purified by IPTG induced Ni2-NTA affinity column. The expression of P5-2 protein was analyzed by SDS-PAGE. [results] the total length of SRBSDV-An Hui-HN2 S5 fragment was 3 167 BP, and the S5-2 gene was 612 BP, encoding 204 amino acids. The results showed that the sequence similarity between SRBSDV-An Hui-HN2 S5 fragment and other SRBSDV isolate S5 fragment was very high. 99.0 and 99.7, and the sequence of S5 fragments of Fijivirus, other members of Fijivirus, is less similar to that of S5 fragments, which are only 38.0% and 71.3%. The phylogenetic tree of S5 fragments constructed shows that SRBSDV and RBSDV are clustered into one branch. Among them, 6 SRBSDV isolates were clustered into one subbranch. Western blot analysis of the recombinant protein with relative molecular weight of 47 000 showed that the McAb could react specifically with the recombinant fusion protein. [conclusion] the SRBSDV isolates have specific reaction with the recombinant fusion protein. [conclusion]. Are very close to each other, The fusion protein expressed in prokaryotic expression was the target protein.
【作者單位】： 安徽農(nóng)業(yè)大學植物保護學院;
【基金】：國家自然科學基金(31371915) 公益性行業(yè)(農(nóng)業(yè))科研專項(201303028)
【分類號】：S435.111.4
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本文編號：1530996