期刊
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
卷 23, 期 16, 页码 -出版社
MDPI
DOI: 10.3390/ijms23169385
关键词
transgenic; Bxb1; integrase; recombinase; GMO
资金
- National Key RD Project [2016YFD0101904]
- Ministry of Science and Technology of China
- Key Research Program of Frontier Sciences CAS, China [QYZDY-SSW-SMC010]
Target sites were generated in the Japonica rice genome using the Mycobacteriophage Bxb1 site-specific integration system. Single copy lines with target sites that were not close to a centromere, known genes or repetitive DNA, and were able to express the reporter gene were successfully produced. New traits were stacked into Japonica rice through gene transfer and site-specific integration.
The clustering of transgenes at a chromosome location minimizes the number of segregating loci that needs to be introgressed to field cultivars. Transgenes could be efficiently stacked through site-specific recombination and a recombinase-mediated in planta gene stacking process was described previously in tobacco based on the Mycobacteriophage Bxb1 site-specific integration system. Since this process requires a recombination site in the genome, this work describes the generation of target sites in the Japonica rice genome. Agrobacterium-mediated gene transfer yielded similar to 4000 random-insertion lines. Seven lines met the criteria of being single copy, not close to a centromere, not inserted within or close to a known gene or repetitive DNA, having precise recombination site sequences on both ends, and able to express the reporter gene. Each target line tested was able to accept the site-specific integration of a new gfp-containing plasmid and in three of those lines, we regenerated fertile plants. These target lines could be used as foundation lines for stacking new traits into Japonica rice.
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