[Objective] The research aimed to isolate flanking sequences adjacent to the transgenic T-DNA in Brassica napus by an improved inverse PCR method.[Method] Using single clone of transgenic FS4 in Brassica napus as the ...[Objective] The research aimed to isolate flanking sequences adjacent to the transgenic T-DNA in Brassica napus by an improved inverse PCR method.[Method] Using single clone of transgenic FS4 in Brassica napus as the research materials,total DNA was extracted from transgenic Brassica napus by using modified CTAB method.After enzyme digestion and purification,self-joining was made.Two circles of nested PCR and the sequence alignment were carried out.[Result] A fragement with the size of 4.0 kb was amplified ...展开更多
Mitogen-activated protein kinases （MAPKs） are a family of Ser/Thr protein kinases widely conserved in all eukaryotes and involved in responses to biotic and abiotic stresses. In this study, two over-expressing BnMAP...Mitogen-activated protein kinases （MAPKs） are a family of Ser/Thr protein kinases widely conserved in all eukaryotes and involved in responses to biotic and abiotic stresses. In this study, two over-expressing BnMAPK1 oilseed rape lines, ov3 and ov11, were used to study the drought-resistant mechanism of BnMAPK1 under natural drought and simulation drought through spraying 10%PEG 8000 in seedlings. Zhongyou 821 （WT） was used as control. Compared with wild type, transgenic seedlings had higher leaf water content, higher root activity, slightly higher peroxidase （POD） and superoxide dismutase （SOD） activity, higher proline content and lower malondialdehyde （MDA） content. The expression of drought-resistant related genes, including P5CSB, PLC, LEA4 and SCE1, have been up-regulated in some degree and the expressed time of transgenic lines were earlier than that of wild type. These results suggested that over-expression of BnMAPK1 can enhance the resistance to drought in oilseed rape （Brassica napus）.展开更多
The seed-specific promoter and transit peptide were amplified and fused to the three genes phbA, phbB and phbC encoding PHB synthetic enzymes, respectively. Seed-specific expression vectors pSCB containing phbC and ph...The seed-specific promoter and transit peptide were amplified and fused to the three genes phbA, phbB and phbC encoding PHB synthetic enzymes, respectively. Seed-specific expression vectors pSCB containing phbC and phbB, and pSCAB containing phbC, phbB and phbA, were constructed by introducing the genes with promoter and peptide into the binary vector pBI101. Transgenic Brassica napus H165 were obtained by Agrobacterium-mediated transformation with these vectors. They were confirmed by PCR, Southern and RT-PCR analyses.展开更多
Integration of a transgene into a C-genome chromosome plays an important role in reducing ecological risk of transgenic Brassica napus To obtain C-genome transgenic B. napus, herbi- cide-resistant bar gene was firstly...Integration of a transgene into a C-genome chromosome plays an important role in reducing ecological risk of transgenic Brassica napus To obtain C-genome transgenic B. napus, herbi- cide-resistant bar gene was firstly transferred into B. oleracea var. alboglabra mediated by Agrobacterium tumefaciens strain LBA4404. Then using the trans- genic B. oleracea as paternal plants and 8 non- transgenic varieties of B. rapa as maternal plants, C- genome transgenic B. napus with bar gene was arti- ficially resynthesized by means of ovary culture and chromosome doubling. Among 67 lines of the resyn- thesized B. napus, 31 were positive, and 36 were negative according to PCR test for bar gene. At least 2 plants from each line were kept for PPT spray con- firmation. The result was in consistence with the PCR test. Genomic Southern blotting of three randomly chosen lines also showed that bar gene had been integrated into the genome of resynthesized B. napus lines.展开更多
基金Supported by National 863 Program of China(2006AA10A113)Natural Science foundation of Zhejiang Province(Y306097)~~
文摘[Objective] The research aimed to isolate flanking sequences adjacent to the transgenic T-DNA in Brassica napus by an improved inverse PCR method.[Method] Using single clone of transgenic FS4 in Brassica napus as the research materials,total DNA was extracted from transgenic Brassica napus by using modified CTAB method.After enzyme digestion and purification,self-joining was made.Two circles of nested PCR and the sequence alignment were carried out.[Result] A fragement with the size of 4.0 kb was amplified ...
基金supported by the National Natural Science Foundation of China (31271756, 31101175)the National "111" Project of China’s Higher Education (B12006)
文摘Mitogen-activated protein kinases （MAPKs） are a family of Ser/Thr protein kinases widely conserved in all eukaryotes and involved in responses to biotic and abiotic stresses. In this study, two over-expressing BnMAPK1 oilseed rape lines, ov3 and ov11, were used to study the drought-resistant mechanism of BnMAPK1 under natural drought and simulation drought through spraying 10%PEG 8000 in seedlings. Zhongyou 821 （WT） was used as control. Compared with wild type, transgenic seedlings had higher leaf water content, higher root activity, slightly higher peroxidase （POD） and superoxide dismutase （SOD） activity, higher proline content and lower malondialdehyde （MDA） content. The expression of drought-resistant related genes, including P5CSB, PLC, LEA4 and SCE1, have been up-regulated in some degree and the expressed time of transgenic lines were earlier than that of wild type. These results suggested that over-expression of BnMAPK1 can enhance the resistance to drought in oilseed rape （Brassica napus）.
文摘The seed-specific promoter and transit peptide were amplified and fused to the three genes phbA, phbB and phbC encoding PHB synthetic enzymes, respectively. Seed-specific expression vectors pSCB containing phbC and phbB, and pSCAB containing phbC, phbB and phbA, were constructed by introducing the genes with promoter and peptide into the binary vector pBI101. Transgenic Brassica napus H165 were obtained by Agrobacterium-mediated transformation with these vectors. They were confirmed by PCR, Southern and RT-PCR analyses.
基金supported by National 863 Project(Grant Nos.2002AA212011,2003AA222101&2005AA241030)the National Natural Science Foundation of China(Grant No.30270791).
文摘Integration of a transgene into a C-genome chromosome plays an important role in reducing ecological risk of transgenic Brassica napus To obtain C-genome transgenic B. napus, herbi- cide-resistant bar gene was firstly transferred into B. oleracea var. alboglabra mediated by Agrobacterium tumefaciens strain LBA4404. Then using the trans- genic B. oleracea as paternal plants and 8 non- transgenic varieties of B. rapa as maternal plants, C- genome transgenic B. napus with bar gene was arti- ficially resynthesized by means of ovary culture and chromosome doubling. Among 67 lines of the resyn- thesized B. napus, 31 were positive, and 36 were negative according to PCR test for bar gene. At least 2 plants from each line were kept for PPT spray con- firmation. The result was in consistence with the PCR test. Genomic Southern blotting of three randomly chosen lines also showed that bar gene had been integrated into the genome of resynthesized B. napus lines.
