薰衣草根际促生菌ACC脱氨酶基因克隆与表达  

Cloning and Expression of ACC Deaminase Gene from Lavender Rhizosphere Growth Promoting Bacteria

作　　者：赵妍妍 林彬 玛依拉·吐尔地别克 恩特马克·布拉提白 ZHAO Yanyan;LIN Bin;MAYILA Tuerdibieke;ENTOMACK Borrathybay(College of Biological Sciences and Technology,Yili Normal University,Yining 835000,China;Xinjiang Key Laboratory of Lavender Conservation and Utilization,Yining 835000,China)

机构地区：[1]伊犁师范大学生物科学与技术学院,新疆伊宁835000 [2]新疆薰衣草资源保护与利用重点实验室,新疆伊宁835000

出　　处：《工业微生物》2024年第4期161-163,共3页Industrial Microbiology

基　　金：国家自然科学基金“伊犁河谷薰衣草根系分泌物对根际促生菌定殖及促生作用的影响”(31960270);国家自然科学基金“红斑丹毒丝菌黏附蛋白SpaA与其受体相互作用及其宿主特异性研究”(3156702)。

摘　　要：本文采用原核表达系统,表达薰衣草根际促生菌的ACC脱氨酶,并探究重组菌酶的ACC脱氨酶活性。经PCR扩增,薰衣草根际促生菌YSX-12的acdS基因长度达到了1 017 bp,与已报道的恶臭假单胞菌AM15株acdS基因核苷酸序列的相似度为99%。构建重组载体p ET-28-ACDS后,在IPTG诱导下,特异表达大小为39 k Da的蛋白条带。Western印迹表明,表达的重组ACC脱氨酶能与小鼠抗6×His标签抗体发生特异性反应;ELISA结果显示,重组ACC脱氨酶能够与抗ACC脱氨酶抗体结合,ACC脱氨酶含量达421 pg/mL,重组菌和YSX-12的最大酶活性分别为0.621 U/mg和0.642 U/mg。通过研究构建的重组菌BL21及其表达的重组ACC脱氨酶,可为进一步研究ACC脱氨酶在根际促生菌中的促生机理奠定基础。This article uses a prokaryotic expression system to express the ACC deaminase of lavender rhizosphere growth promoting bacteria,and explore the ACC deaminase activity of recombinant bacterial enzymes.After the PCR amplification,the length of the acdS gene of lavender rhizosphere growth promoting bacterium YSX-12 was 1017 bp,and the nucleotide sequence similarity of it with the reported the acdS gene of pseudomonas putida AM15 strain was 99%.After constructing the recombinant vector pET-28-ACDS,a protein band of 39 kDa was specifically expressed under the IPTG induction.Western blotting showed that the expressed recombinant ACC deaminase could react specifically with mouse anti 6×His tag antibodies.The ELISA results showed that recombinant ACC deaminase could bind to anti ACC deaminase antibodies,with an ACC deaminase content of 421pg/mL,and the maximum enzyme activity of recombinant bacteria and YSX-12 were respectively 0.621 U/mg and 0.642 U/mg,respectively.By studying the constructed recombinant bacterial strain BL21 and its expressed recombinant ACC deaminase,a foundation is laid for further studying the mechanism of ACC deaminase in promoting rhizosphere growth of bacteria.

关 键 词：根际促生菌 ACC脱氨酶 基因克隆 原核表达 酶活性 

分 类 号：Q78[生物学—分子生物学] Q936