兰州大学机构库 >生命科学学院
The guanidine thiocyanate-high EDTA method for total microbial RNA extraction from severely heavy metal-contaminated soils
Pei, Yaxin1,2; Mamtimin, Tursunay1,2; Ji, Jing1,3; Khan, Aman1,3; Kakade, Apurva1,2; Zhou, Tuoyu1,3; Yu, Zhengsheng2,3; Zain, Hajira1,3; Yang, Wenzhi1; Ling, ZM(令桢民)1,2,3; Zhang, WY(张文雅)1; Zhang, YM(张迎梅)1; Li, XK(李祥锴)1,2,3
2020-06-23
Source PublicationMICROBIAL BIOTECHNOLOGY
ISSN1751-7915
AbstractMolecular analyses relying on RNA, as a direct way to unravel active microbes and their functional genes, have received increasing attention from environmental researchers recently. However, extracting sufficient and high-quality total microbial RNA from seriously heavy metal-contaminated soils is still a challenge. In this study, the guanidine thiocyanate-high EDTA (GTHE) method was established and optimized for recovering high quantity and quality of RNA from long-term heavy metal-contaminated soils. Due to the low microbial biomass in the soils, we combined multiple strong denaturants and intense mechanical lysis to break cells for increasing RNA yields. To minimize RNAase and heavy metals interference on RNA integrity, the concentrations of guanidine thiocyanate and EDTA were increased from 0.5 to 0.625 ml g(-1)soil and 10 to 100 mM, respectively. This optimized GTHE method was applied to seven severely contaminated soils, and the RNA recovery efficiencies were 2.80 similar to 59.41 mu g g(-1)soil. The total microbial RNA of non-Cr(VI) (NT) and Cr(VI)-treated (CT) samples was utilized for molecular analyses. The result of qRT-PCR demonstrated that the expressions of two tested genes,chrAandyieF, were respectively upregulated 4.12- and 62.43-fold after Cr(VI) treatment. The total microbial RNA extracted from NT and CT samples, respectively, reached to 26.70 mu g and 30.75 mu g, which were much higher than the required amount (5 mu g) for metatranscriptomic library construction. Besides, ratios of mRNA read were more than 86%, which indicated the high-quality libraries constructed for metatranscriptomic analysis. In summary, the GTHE method is useful to study microbes of contaminated habitats.
DOI10.1111/1751-7915.13615
Indexed BySCIE
Language英语
Funding ProjectNational Natural Science Foundation[31870082] ; Gansu Province Major Science and Technology Projects[17ZD2WA017] ; Fundamental Research Funds for the Central Universities[lzujbky-2018-it47]
WOS Research AreaBiotechnology & Applied Microbiology ; Microbiology
WOS SubjectBiotechnology & Applied Microbiology ; Microbiology
WOS IDWOS:000542034200001
PublisherWILEY
Original Document TypeArticle ; Early Access
Citation statistics
Cited Times:1[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.lzu.edu.cn/handle/262010/405735
Collection生命科学学院
Corresponding AuthorLi, Xiangkai
Affiliation1.Lanzhou Univ, Gansu Key Lab Biomonitoring & Bioremediat Environ, Sch Life Sci, Tianshuinanlu 222, Lanzhou 730000, Gansu, Peoples R China
2.Lanzhou Univ, Ministry Educ, Key Lab Cell Act & Stress Adaptat, Sch Life Sci, Tianshuinanlu 222, Lanzhou 730000, Gansu, Peoples R China
3.Gansu Acad Membrane Sci & Technol, Key Lab Resources Utilizat Technol Unconvent Wate, Duanjiatanlu 1272, Lanzhou 730000, Gansu, Peoples R China
First Author AffilicationSchool of Life Sciences
Corresponding Author AffilicationSchool of Life Sciences
Recommended Citation
GB/T 7714
Pei, Yaxin,Mamtimin, Tursunay,Ji, Jing,et al. The guanidine thiocyanate-high EDTA method for total microbial RNA extraction from severely heavy metal-contaminated soils[J]. MICROBIAL BIOTECHNOLOGY,2020.
APA Pei, Yaxin.,Mamtimin, Tursunay.,Ji, Jing.,Khan, Aman.,Kakade, Apurva.,...&Li, Xiangkai.(2020).The guanidine thiocyanate-high EDTA method for total microbial RNA extraction from severely heavy metal-contaminated soils.MICROBIAL BIOTECHNOLOGY.
MLA Pei, Yaxin,et al."The guanidine thiocyanate-high EDTA method for total microbial RNA extraction from severely heavy metal-contaminated soils".MICROBIAL BIOTECHNOLOGY (2020).
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