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Abstract All experiment methods were performed according to the standard of molecular clone technology. The DNA fragment of GFP gene was inserted into the pBI121-Lyz to get the recombined eukaryotic expression vector pBI121-Lyz-GFP. Polymerase chain reaction (PCR) was used to proliferate GFP gene from pBI121-Lyz-GFP plasmid, producing an approximate 750bp band with restriction enzyme SmaI and BamHI at both sides of the DNA. The eukaryotic expression vector was also verified by nucleotide sequencing. Plasmid pBI121-Lyz-GFP was introduced into agrobacterium tumenfaciens strain LBA4404. Discs of HeTian alfalfa were transformed with LBA4404 strain. Kanamycin resistance was selected and induced to regenerate alfalfa callus. The observation of GFP expression in HeTian alfalfa callus with slides was carried out under the fluorescent microscope upon excitation of blue light. PCR amplification and the results of observation showed that the recombined plasmid was transformed into HeTian alfalfa callus successfully.
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Received: 17 December 2007
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