[Cloning of UGT gene of Bupleurum chinense and construction of over expressing and RNAi transgenic vectors].
OBJECTIVE
To clone the full-length cDNA of a uridine diphosphate glycosyltransferase (UGT) gene which may be involved in saikosaponin biosynthesis of Bupleurum chinense, and construct the transgenic vectors for over expression and RNAi of the cloned UGT. These works will provide foundation for further its function analysis by transgene study.


METHODS
RAGE and LD-PCR were used to clone the full-length cDNA of the UGT, on the basis of its partial cDNA sequence obtained from our previous 454-sequenced dataset. The ORF and partial sequences of the UGT were PCR cloned using primers with corresponding restriction enzymes cutting sites. The PCR products were digested with corresponding restriction enzymes and then were inserted into pCAMBIA-SUPER 1 300 and pHANNIBAL. The recombinant pHANNIBAL were digested with Not I and then were inserted into a binary vector, pART27. Finally, the transgenic vectors for over expression and RNAi of the cloned UGT were constructed.


RESULTS
The full-length cDNA of a UGT were cloned from B. chinense. The recombinant vectors for over expression and RNAi of the UGT were obtained.


CONCLUSIONS
Our works on full-length cDNA cloning and transgenic vectors construction provide a substantial foundation for follow-up biofunction analysis of the UGT through transgenic research.
Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100193, China. csui@implad.ac.cn
Chun
C
Sui
Jiesen
J
Xu
Lizi
L
Zhao
Jianhe
J
Wei
Yanhong
Y
Xu
Peng
P
Sun
D000595:Amino Acid Sequence; D029972:Bupleurum / Q000235:genetics*; D003001:Cloning, Molecular; D005822:Genetic Vectors; D014453:Glucuronosyltransferase / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D034622:RNA Interference; D019076:Transgenes
D014453:Glucuronosyltransferase