刺五加鲨烯环氧酶基因cDNA的克隆及序列分析  

Cloning and sequence analysis on cDNA of squalene epoxidase gene in Eleutherococcus senticosus

作　　者：邢朝斌[1] 曹蕾[1] 陈龙[1] 何闪[1] 李宝财[1] 朱金丽[1] 

机构地区：[1]河北联合大学生命科学学院,河北唐山063000

出　　处：《中国中药杂志》2012年第2期172-175,共4页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金项目(30701086);河北省自然科学基金项目(C2009001252)

摘　　要：目的:对刺五加鲨烯环氧酶基因的cDNA进行克隆及序列分析。方法:采用改良的异硫氰酸胍法提取刺五加总RNA,逆转录为cDNA,根据已报道的人参SE基因cDNA序列设计引物,利用RT-PCR法克隆刺五加SE基因的cDNA序列。结果:克隆了2个序列不同的cDNA(SE1和SE2),开放阅读框分别长1 665,1 629 bp,分别编码554,542个氨基酸。SE1,SE2间的核苷酸和氨基酸一致性分别为91.49%,92.55%,两者与三七SE1的氨基酸序列相似性最高,分别为93.45%,94.87%。SE1,SE2均含有1个FAD结合区域,SE1,SE2推测的氨基酸分别存在2个和4个跨膜螺旋。结论:首次分离并报道了刺五加的2个SE基因cDNA序列,为刺五加的次生代谢工程研究奠定了基础。Objective:To clone and sequence the cDNA of squalene epoxidase gene in Eleutherococcus senticosus.Method:Total RNA of E.senticosus was extracted by the improved isothiocyanate method and reverse transcripted into cDNA.The primers were designed depending on the reported SE cDNA sequences of Panax ginseng.The SE cDNAs in E.senticosus was amplified using RT-PCR strategy.Result:Sequencing results showed two different cDNA fragments(SE1,SE2) with 1 665,1 629 bp each ORF which encoded 554,542 amino acids,respectively.The identities of nucleotides and amino acids between SE1,SE2 were 91.49%,92.55%.SE1,SE2 had the highest amino acids similarity to the SE1 of P.notoginseng,93.45%,94.87% respectively.SE1,SE2 both had a FAD binding domain.The deduced speculated amino acids of SE1,SE2 each had 2,4 membrane-spanning helices.Conclusion:The two SE sequences in E.senticosus were firstly separated and reported,which has made foundation for E.senticosus secondary metabolite engineering researches.

关 键 词：刺五加 鲨烯环氧酶基因 克隆 RT-PCR 

分 类 号：R282.5[医药卫生—中药学]