摘要
为克隆欧李钙调蛋白基因序列,本研究采用CODEHOP方法设计了一对简并引物,F1:5'-GGCTGACCAACTGACCga(c/t)ga(c/t)ca(a/g)at-3';F2:5'-CACGTCAGCCTCTCTGatcat(c/t)tc(a/g)tc-3';并采用传统简并引物设计方法设计了简并引物N1:5’-AAATGGC(A/C/G)GATCAGCT(A/C/T)AC-3’;N2:5:’-CACTT(A/G)GCCATCATGAC(C/T)TT-3’.从欧李的幼苗组织中成功克隆出了钙调蛋白基因片段,并第一次完成了对该序列的测序.通过利用NCBI的Blast进行分析等方法证实了该序列与其他植物的钙调蛋白序列具有高度的同源性,该序列与梅花、桃、甜樱桃的钙调蛋白序列同源性都在90%以上.研究证明CODEHOP软件相比传统的设计简并引物的方法具有可信性和高效性,并且钙调蛋白目的基因的成功克隆对钙调蛋白以及欧李高钙特性的研究提供了基础数据.
In this study, a pair of degenerate primers, F1 : 5 '-GGCTGACCAACTGACCga (c/t) ga (c/t) ca (a/g) at-3 : F2:5-CACGTCAGCCTCTCTGatcat (c/t) tc (a/g) tc-3', are designed by CODEHOP. And degenerate primers, N1 : 5' -AAATGGC(A/C/G) GAT CAGCT(A/C/T) AC-3', N2 : 5' -CACTT(A/G) GCCATCATGAC (C/T) 33'-3', are designed by traditional strategy. Two CaM fragments are amplified from seedling of Cerasus humilis, and the calmodulin gene fragment is sequenced the first time. Blast analysis on NCBI, shows that the sequence has high ho- mology with other calmodulin genes. Compared with Prunus mume, Prunus persica and Prunus avium, the homology rates are more than 90%. The results also suggeste that a degenerate primer designed by CODEHOP is credibility and efficient. Successfully cloning the CaM fragment provied basic information for the research of calmoulin and high-calcium characteristic of Cerasus humilis.
作者
邬晓勇
孙雁霞
梁勇
陈月星
苟小军
WU Xiao-yong SUN Yan-xia LIANG Yong CHEN Yue-xing GOU Xiao-jun(School of Pharmacy and Bioengineering, Chengdu University, Chengdu 610106, China)
出处
《烟台大学学报(自然科学与工程版)》
CAS
2017年第4期292-300,共9页
Journal of Yantai University(Natural Science and Engineering Edition)
基金
教育部地方高校国家级大学生创新创业训练计划项目(201411079017)
成都大学大学生创新试验计划(CDUCX2015108)