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学科主题物理化学
High-Quality RNA Preparation from Rhodosporidium toruloides and cDNA Library Construction Therewith
Yang, Fan1,2; Tan, Haidong1; Zhou, Yongjin1,2; Lin, Xinping1,2; Zhang, Sufang1; Zhang SF(张素芳)
关键词Rna Extraction Rhodosporidium Toruloides Full-length Cdna Library
刊名MOLECULAR BIOTECHNOLOGY
2011-02-01
ISSN待补充
DOI10.1007/s12033-010-9322-1
47期:2页:144-151
收录类别SCI
文章类型Article
部门归属18
项目归属1816
产权排名1,1
WOS标题词Science & Technology ; Life Sciences & Biomedicine
类目[WOS]Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
研究领域[WOS]Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
关键词[WOS]LIPID-ACCUMULATION ; OLEAGINOUS MICROORGANISMS ; PLANT-TISSUES ; EXTRACTION ; RICH ; POLYSACCHARIDES ; PROTOCOL ; CULTURE ; BIOLOGY
英文摘要High-Quality RNA Preparation from Rhodosporidium toruloides and cDNA Library Construction Therewith; Oleaginous yeast Rhodosporidium toruloides is an excellent microbial lipid producer. Therefore, it is important to develop molecular biology tools to understand the basic mechanism for lipid accumulation and further manipulate the microorganism. High-quality RNA extraction from R. toruloides is particularly challenging due to high level of polysaccharides, lipids, and other secondary metabolites. To obtain an optimal protocol for RNA extraction from R. toruloides, four methods were evaluated. Large difference in RNA yield and quality among these protocols was found. The optimum method was modified RNAiso procedure, where RNA was isolated using liquid nitrogen-RNAiso method with salt precipitation and the addition of beta-mercaptoethanol. This method consistently recovered RNA in good quality with high yield. Around 297 mu g total RNA per gram of cells was obtained with an average purity measured as A(260)/A(280) of 2.09. A titer of 10(5) cfu/ml could be harvested to construct a full-length cDNA library with the RNA sample in this quality. Electrophoresis gel analysis indicated the fragments ranged from 200 bp to 4.0 kb, with the average size of 1000 bp. Randomly picked clones showed the recombination efficiency at 80%. These results showed that RNA of R. toruloides was successfully extracted for the first time using the modified RNAiso method, and the cDNA library was appropriate for screening the genes related to lipid accumulation.
语种英语
WOS记录号WOS:000286378600006
引用统计
文献类型期刊论文
条目标识符http://cas-ir.dicp.ac.cn/handle/321008/115574
专题中国科学院大连化学物理研究所
通讯作者Zhang SF(张素芳)
作者单位1.CAS, Dalian Inst Chem Phys, Div Biotechnol, Dalian 116023, Peoples R China
2.Chinese Acad Sci, Grad Univ, Beijing 100049, Peoples R China
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GB/T 7714
Yang, Fan,Tan, Haidong,Zhou, Yongjin,et al. High-Quality RNA Preparation from Rhodosporidium toruloides and cDNA Library Construction Therewith[J]. MOLECULAR BIOTECHNOLOGY,2011,47(2):144-151.
APA Yang, Fan,Tan, Haidong,Zhou, Yongjin,Lin, Xinping,Zhang, Sufang,&张素芳.(2011).High-Quality RNA Preparation from Rhodosporidium toruloides and cDNA Library Construction Therewith.MOLECULAR BIOTECHNOLOGY,47(2),144-151.
MLA Yang, Fan,et al."High-Quality RNA Preparation from Rhodosporidium toruloides and cDNA Library Construction Therewith".MOLECULAR BIOTECHNOLOGY 47.2(2011):144-151.
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