Construction of subtractive cDNA library of genes involved in saponin biosynthesis from different age sanchi root | 不同年龄三七根皂甙合成相关基因的差减cDNA文库构建
2010
He Fengmei, Yunnan Agricultural University, Kunming(China), College of Horticulture and Landscape | Zhu Yongping, Yunnan Agricultural University, Kunming(China), College of Agriculture and Biotechnology
Chinese. 本研究以三七3年牛根和1年生根为材料，利用抑制消减杂交技术构建三七根在不同年龄阶段皂甙合成相关基因的差减cDNA文库，正向文库（3年生文库）和反向文库（1年生文库）分别包括2000个和500个cDNA克隆，重组率在95％以上；外源片段的长度分布在200～900bp，平均片段长度约500bp；差减效率检测表明：消减前后基因的拷贝数相差32000倍，差异表达的基因得到了充分的富集。结果表明所构建的cDNA文库符合SSH文库的质量标准，能够满足下一步的实验要求，为研究差异表达基因，探讨皂甙生物合成的分子机理，提高三七皂甙产量产量和质量奠定基础。[著者文摘]
Show more [+] Less [-]English. The roots of 3-year-old and 1-year-old sanchi were used to be as experiment materials, the subtractive cDNA library of genes related to saponin biosynthesis from different age sanchi root were constructed by using suppression subtractive hybridization in this study. The forward-subtracted cDNA library of 3-year-old root had 2 000 clones and reverse library of 1-year-old root had 500 clones, respectively, The recombination fraction of the both subtractive libraries above 95%, the inserted fragments between 200 bp and 900 bp with average fragment size was 500 bp. The subtractive efficiency showed that copy number ofgene before and after subtractive hybridization was 32 000 fold, and differential expression of genes get a remarkable enrichment. Further results indicated that the cDNA library quality are exactly within SSH library standard and could be used in further studies, also would provide reference for studying the differentially expressed genes to explore the molecular mechanism of saponin biosynthesis, and increase productivity and quality of saponins content in sanchi.[著者文摘]
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