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Construction of
full-length
cDNA library of white flower
Salvia miltiorrhiza bge f.alba root and partial EST
sequence analysis
Gangping Hao*, Renjiu Shi,
Jianmei Wang* and Bing Qi
*Department
of Biological Science, Taishan Medical University, Taian
271000, China.
*Corresponding author. E-mail:
haogangping@163.com,
jmwang@tsmc.edu.cn
Tel: +86-538-6222043. Fax: +86-538-6222053.
Accepted
2 April, 2009 |
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In order to
screen and isolate secondary metabolite biosynthesis related
gene, we construct a cDNA library of white flower
Salvia miltiorrhiza bge. f.alba.
High quality of total RNA was successfully isolated from
roots of white flower
S. miltiorrhiza
using
modified CTAB method. Double strand cDNA was cloned into
pDNR-LIB
vector. The number of clones,
recombinant rate and length of insert fragments were
determined. Results showed that the capacity of the original
library was 1.8×107 with a recombinant rate of 91%
and the inserted cDNA fragments ranged from 0.5 to 2.0 kb.
Partial cDNAs chosen by random were sequenced. After
BLAST analysis of some cDNAs, their possible functions were
predicted. It is found that most of these cDNAs were similar
to homological genes of
Arabidopsis thaliana, Oryza sativa, and other
plants. Most of the genes were related to cell metabolism,
stress resistance, cell growth and development, etc. More
importantly, some key enzymes and factors involved in
secondary metabolism of S. miltiorrhiza,
such as EST fragments of phenylalanine ammonialyase (SmPAL1),
chorismate synthase (SmCHS),
3-hydroxy-3-methylglutaryl CoA reductase (SmHMGR),
4-Coumarate-coenzyme A ligase (Sm4CL1) and SmMYB90,
were found from this library. These results indicated that
the library has enough capacity, high recombinant rate and
long insert fragment. This study provided a base for
further study on the structure and function of these cDNAs.
Key
words:
white flower Salvia
miltiorrhiza bge f.alba, RNA isolation, cDNA library
construction, EST sequence analysis. |
