Construction of Subtractive cDNA Library from the Leaves of Dendrobium candidum Wall, ex Lindl. by Suppression Subtractive Hybridization Technology
- VernacularTitle:应用抑制性差减杂交技术构建铁皮石斛差减cDNA文库的探讨
- Author:
Xiaoyong WEI
- Publication Type:Journal Article
- Keywords:
DENDROBINE /biosynthesis;
SUBTRACTIVE CDNA LIBRARY;
SUPPRESSION SUBTRACTIVE HYBRIDIZATION;
DENDROBIUM CANDIDUM
- From:
Journal of Guangzhou University of Traditional Chinese Medicine
2000;0(04):-
- CountryChina
- Language:Chinese
-
Abstract:
[Objective] Subtractive cDNA libraries from the leaves of D. candidum were constructed to supply evidence for the clone of dendrobine-biosynthesis-associated genes. [ Methods ] Suppression subtractive hybridization ( SSH) technology was used. With the cDNA of perennial leaf as the tester and that of annual leaf as the driver, forward and reverse hybridization was performed. The two obtained subtracted cDNA fragments were cloned into PointTMXa-1 plasmid vectors, and then the vectors were transformed into E. Coli JM109. The inserted fragments were amplified by PCR and then identified. [Results] The subtractive cDNA libraries related with dendrobine-biosynthesis-associated genes were successfully constructed. The forward and reverse subtractive libraries included 560 and 220 clones respectively. Detected by PCR, the length of the inserted fragments was 550 bp on average. [Conclusion] The constructed subtractive libraries are suitable for further study on the functional genes associated with dendrobine biosynthesis of D. candidum.
