The construction of an Agrobacterium-mediated transformation system of Gynostemma pentaphyllum using the phosphomannose-isomerase/mannose selection system.
IF 1.4 4区 生物学Q4 BIOTECHNOLOGY & APPLIED MICROBIOLOGY
Muxiu Tan, Fengming Liu, Yueying Xie, Qiaocheng Mo, Fenghua Shi
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引用次数: 0
Abstract
In this study, the transformed system mediated by Agrobacterium tumefaciens of Gynostemma pentaphyllum was constructed by using the phosphomannose-isomerase (PMI) gene as a marker. To investigate the cefotaxime sodium salt (Cef) concentration of bacteriostatic medium and the appropriate mannose concentration in the selectable medium, explants of the stems with buds were cultured in a basic medium supplemented with different Cef and mannose concentrations, respectively. After these were optimized, 288 explants were transformed according the protocol described above to verify their availability by using the polymerase chain reaction (PCR), reverse transcription-PCR and chlorophenol red. The results showed that the appropriate Cef concentration for bacteriostatic culture and mannose concentration for selectable culture were 150 mg l-1 and 3 g l-1 for stem with buds, respectively. According to the PCR results, the transformation frequency of stems with buds was 20.49% with a regeneration rate of 29.16%. In future, the CPR assay could be the auxiliary method of choice as it is moderately accurate, but it has good maneuverability and is cost effective for large-scale use.
期刊介绍:
Plant Biotechnology is an international, open-access, and online journal, published every three months by the Japanese Society for Plant Biotechnology. The journal, first published in 1984 as the predecessor journal, “Plant Tissue Culture Letters” and became its present form in 1997 when the society name was renamed to Japanese Society for Plant Cell and Molecular Biology, publishes findings in the areas from basic- to application research of plant biotechnology. The aim of Plant Biotechnology is to publish original and high-impact papers, in the most rapid turnaround time for reviewing, on the plant biotechnology including tissue culture, production of specialized metabolites, transgenic technology, and genome editing technology, and also on the related research fields including molecular biology, cell biology, genetics, plant breeding, plant physiology and biochemistry, metabolic engineering, synthetic biology, and bioinformatics.