欧美杨107木质素生物合成酶CCR基因的克隆及其鉴定  

Cloning and Identification of Lignin Biosynthesis CCR Gene from Populus×euramericana cv."74/76"

作　　者：薛永常[1] 赵一玲[1] 赵文超[1] 王雪霞[1] 

机构地区：[1]大连工业大学生物学院辽宁省发酵工程重点实验室,辽宁大连116034

出　　处：《辽宁林业科技》2008年第5期18-21,30,共5页Liaoning Forestry Science and Technology

基　　金：辽宁省自然科学基金(20062146);辽宁省教育厅科研项目(20040091)

摘　　要：木质素作为草类饲料消化性的限制因子,直接影响饲料的利用率,而CCR是木质素生物合成关键酶之一。从正在分化的2a生欧美杨107次生木质部RT-PCR扩增出的基因片段,与pMD20-T载体连接,重组质粒经限制性内切酶酶切、特异引物PCR扩增和测序鉴定。结果表明该扩增片段长为961bp,含一个编码301氨基酸的完整开放阅读框,其核苷酸序列与Leple等发表的白杨杂种Populus trichocarpa CCR的cDNA序列(AJ224986)同源性达到98.2%,并且反向插入到pMD20-T载体,因此构建了CCR的反义大肠杆菌表达载体,为后续的基因操作打下基础。该基因序列已提交国际核苷酸序列库,登录号为AM921698。Lignin is the limiting factor of forage plants＇ feeding value and cell wall digestibility. Cinnamoyl- CoA reductase is one important lignin synthetase. In this paper, a CCR fragment was amplified by RT- PCR from poplar（ Populus × euramer/cana cv. ＂74/76＂） developing second xylem mRNA, and be cloned into pMD20- T vector, then identified by restriction enzyme, PCR amplification and sequencing. The sequence of the amplified DNA fragment was 961 base pairs and contained an Oil＂ which translate a protein with 301 amino acids. And it was cloned into pMD20- T vector in reverse direction. Alignment with eDNA sequence of CCR retrieved from NCBI nucleotide database（accession number AJ224986） showed that their homology was 98.2%. So the cloned DNA fragment could be part of CCR. So we constructed the antisense expressional vector for CCR in E. Coli. It was submitted to Genbank, the accession number was AM921698.

关 键 词：CCR RT—PCR 克隆 木质素 

分 类 号：Q946.59[生物学—植物学]