Construction of plant expression vectors with PMI gene as selection marker and their utilization in transformation of Salvia miltiorrhiza f. alba.
- Author:
Ru TAO
;
You-Can ZHANG
;
Qian FANG
;
Ren-Jiu SHI
;
Yan-Ling LI
;
Lu-Qi HUANG
;
Gang-Ping HAO
- Publication Type:Journal Article
- MeSH: Anti-Bacterial Agents; pharmacology; Biomarkers; Cinnamates; pharmacology; Escherichia coli; enzymology; genetics; Escherichia coli Proteins; genetics; metabolism; Gene Expression; Genetic Vectors; genetics; metabolism; Hygromycin B; analogs & derivatives; pharmacology; Mannose-6-Phosphate Isomerase; genetics; metabolism; Plants, Genetically Modified; drug effects; genetics; metabolism; Salvia miltiorrhiza; drug effects; genetics; metabolism; Transformation, Genetic
- From: China Journal of Chinese Materia Medica 2014;39(7):1209-1213
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct plant expression pCAMBIA1301-PMI by substituting PMI for hygromycin resistance gene in pCAMBIA1301 and obtain transgenic Salvia miltiorrhiza f. alba using PMI-mannose selection system.
METHODThe 6-phosphomannose isomerase gene (PMI) of Escherichia coli was amplified by PCR. Sequence analysis showed that it shared 100% amino acids identities with the sequences of PMI genes isolates reported in the NCBI. Based on pCAMBIA1305, the plant expression pCAMBIA1305-PMI was constructed successfully by substituting PMI for hygromycin resistance gene in pCAMBIA1305. pCAMBIA1305-PMI was transformed into Agrobacterium tumefaciens LBA4404, and then the leaves of S. miltiorrhiza f. alba were inoculated in LBA4404 with pCAMBIA1305-PMI.
RESULTPlant expression pCAMBIA1301-PMI was successfully constructed and the leaves of S. miltiorrhiza f. alba inoculated in LBA4404 with pCAMBIA1305-PMI were selected on medium supplemented with a combination of 20 g x L(-1) mannose and 10 g x L(-1) sucrose as a carbon source. The transformation efficiency rate was 23.7%.
CONCLUSIONGenetic transformation was confirmed by PCR, indicating that a new method for obtaining transgenic S. miltiorrhiza f. alba plants was developed using PMI-mannose selection system.