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人参皂苷生物合成相关新基因GBR6的cDNA克隆及序列分析 被引量:3
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作者 刘水平 罗志勇 +4 位作者 陈湘晖 文斌 汪凌昊 魏来 胡维新 《生命科学研究》 CAS CSCD 2004年第4期351-354,共4页
从人参根组织中分离总RNA,使用RT-PCR扩增出人参皂苷生物合成相关基因GBR6开放阅读框(ORF)cDNA片断,并将其定向克隆入原核高效表达载体pQE30,阳性克隆经BamHⅠ和PstⅠ双酶切鉴定、测序验证与已知的GBR6ORF序列完全一致.因而成功构建了pQ... 从人参根组织中分离总RNA,使用RT-PCR扩增出人参皂苷生物合成相关基因GBR6开放阅读框(ORF)cDNA片断,并将其定向克隆入原核高效表达载体pQE30,阳性克隆经BamHⅠ和PstⅠ双酶切鉴定、测序验证与已知的GBR6ORF序列完全一致.因而成功构建了pQE30-GBR6ORF融合原核表达载体,为下一步进行GBR6功能表达分析奠定了基础. 展开更多
关键词 人参皂苷生物合成 gbr6基因 CDNA克隆 序列分析
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人参植物高质量RNA的分离及其皂苷生物合成相关新基因GBR6的表达分析 被引量:4
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作者 罗志勇 刘水平 +5 位作者 陈湘晖 文斌 陆秋恒 罗建清 熊德慧 胡维新 《生命科学研究》 CAS CSCD 2004年第1期52-57,共6页
为从富含多糖的人参植物组织中分离出高质量的RNA,使用改进的两种异硫氰酸胍法,可从人参植物根组织中提取到较高产量和质量的总RNA。每克新鲜组织的RNA产量在80~110μg之间;经琼脂糖凝胶电泳分析,可见到28S和18SrRNA两条完整、清晰的条... 为从富含多糖的人参植物组织中分离出高质量的RNA,使用改进的两种异硫氰酸胍法,可从人参植物根组织中提取到较高产量和质量的总RNA。每克新鲜组织的RNA产量在80~110μg之间;经琼脂糖凝胶电泳分析,可见到28S和18SrRNA两条完整、清晰的条带;紫外分光光度法测得的A_(260)/A_(280)比值位于1.8~2.0。而且,使用该改良法分离的RNA,可广泛用于检测基因表达的RT-PCR与Northern印迹杂交分析以及cDNA文库构建等植物分子生物学研究。 展开更多
关键词 RNA 分离 皂苷 生物合成 基因 gbr6 基因表达 异硫氰酸胍法 人参植物
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Spatio-Temporal Expression Pattern of Six Novel Candidate Genes in Ginsenoside Biosynthesis from Panax ginseng C. A. Meyer 被引量:1
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作者 Zhi-YongLUO Shui-PingLIU +5 位作者 Xiang-HuiCHEN YingRUAN Jian-QingLUO BinWEN Chun-LinLIU Wei-XinHU 《Journal of Integrative Plant Biology》 SCIE CAS CSCD 2005年第5期622-631,共10页
Abstract: To explore the mode of the spatio-temporal expression of six newly discovered ginsenoside biosynthesis candidate gene transcripts, both Northern blotting and semi-quantitative reverse transcription-polymeras... Abstract: To explore the mode of the spatio-temporal expression of six newly discovered ginsenoside biosynthesis candidate gene transcripts, both Northern blotting and semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) were used to elucidate the mRNA expression levels of the transcripts in various tissues and organs of Panax ginseng C. A. Meyer during different growth development stages. The six gene transcripts were all differentially expressed in cultured callus, root, stem, leaf, and seed. The mRNA expression levels were significantly higher in four-year-old roots than in one-year-old roots, and results of semi-quantitative RT-PCR assays were in accordance with those of Northern blotting analyses. The results strongly suggest that all six genes were differentially expressed at root-specific developmental stages. In particular, when a quiescent early stage culture suspension of P. ginseng cells was exposed to the ginsenoside biosynthesis-promoting elicitor Aspergillus niger polysaccharide, the GBR6 gene transcript response showed time-dependent increments and was parallel with ginsenoside productivity (P < 0.01). Overexpressionof the GBR6 gene is likely to play a critically important role in the biosynthesis of ginsenosides. The results of the present study provided a background for the further elucidation of the structure and physiological function of these six candidate genes. 展开更多
关键词 Aspergillus niger polysaccharide ELICITOR gbr6 mRNA expression ginsenoside biosynthesis ginsenoside biosynthesis candidate genes mRNA differential expression Panax ginseng C. A. Meyer
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