摘要
从采绒革盖菌(Coriolus versicolor,常被称为杂色云芝)中筛选到产植酸酶基因.根据植酸酶基因的保守序列设计引物P1、P2,从云芝中分离克隆植酸酶基因的片段.再在分离到的已知序列的5′端和3′端设计嵌套的特异引物SP1、SP2、SP3和SP1′、SP2′、SP3′,利用TAIL-PCR技术进行分离已知序列的侧翼序列.结合Blastn、ORF和Primer5.0软件,在侧翼序列上设计特异引物P3、P4,最终从云芝中分离克隆到全长的植酸酶基因B4.再在B4序列的基础上设计引物phyP rt1、phyP rt2,利用这两个引物通过RT-PCR技术从云芝中克隆到全长的植酸酶基因B5.通过分析,分离克隆到的植酸酶基因可能是新的基因.
Phytase gene was screened from Coriolus versicolor in this study. P1, P2 were designed according to the consensus sequence of phytase gene. Phytase gene fragment was isolated by primers P1, 1）2. The nested special primers, SP1, SP2, SP3 and SPI＇, SP2＇, SP3＇ on the 5＇-flanking and 3＇-flanking on the known sequence were designed, and then the flanking sequences of known fragment were isolated by TAIL-PCR. These sequences were analyzed with blastn, ORF and Primer 5.0 softwares, special primers P3, P4 on the flanking sequence were designed. Finally, the whole phytase gene sequence P,4 was cloned by the primers P3, P4 from C. versicolor. Primers phyP rtl, phyP rt2 were designed on basis of the sequence of B4, then another phytase gene sequence B5 was cloned by RT-PCR through those two primers. After analyzed by some softwares, the isolated gene sequence may be a new phytase gene.
出处
《应用与环境生物学报》
CAS
CSCD
北大核心
2006年第5期618-622,共5页
Chinese Journal of Applied and Environmental Biology
基金
国家自然科学基金资助项目(No.30060054
No.30371000)
广东省自然科学基金资助项目(No.032239)
华南农业大学校长基金项目(5100-k03003)的部分内容~~
