茶树低温胁迫下microRNA实时定量PCR内参基因的筛选

doi:10.13305/j.cnki.jts.2015.06.013

Abstract

Abstract: MicroRNA (miRNA) plays an important role in post transcriptional regulation of gene expression. The research of miRNA quantitative expression level has become more and more popular. Choosing a suitable reference gene is a prerequisite of an accurate real-time fluorescence quantitative PCR(qRT-PCR) assay. The expressions of 9 candidate reference genes by qRT-PCR method in different tea (Camellia sinensis) samples were investigated and their stabilities were analyzed by BestKeeper and geNorm. The results showed that a novel tea miRNA (PC-3p-222) was the most stable reference gene in different tea plant tissues under different cold temperature. The average Ct value was 22-23 which showed moderate expression abundance. Therefore, it can be used as the appropriate reference gene for miRNA qRT-PCR under cold stress, which provides a good choice for the quantitative of miRNA expression.

Key words: Camellia sinensis, miRNA, reference gene, qRT-PCR

CLC Number:

S571.1
Q52

XIE Xiaofang, TIAN Xianfeng, JIANG Changjun, LI Yeyun. Screening of microRNA Reference Genes for Real-time Fluorescence Quantitative PCR under Cold Stress in Camellia sinensis[J]. Journal of Tea Science, 2015, 35(6): 596-604.

References 24

[1]	Heid CA, Stevens J, Livak KJ, et al. Real time quantitative PCR[J]. Genome Res, 1996, 6(10): 986-994.
[2]	Bustin SA, Benes V, Nolan T, et al. Quantitative real-time RT-PCR-a perspective[J]. J Mol Endocrinol, 2005, 34(3): 597-601.
[3]	Ginzinger DG.Gene quantification using real-time quantitative PCR: an emerging technology hits the mainstream[J]. Exp Hematol, 2002, 30(6): 503-512.
[4]	Klein D.Quantification using real-time PCR technology: applications and limitations[J]. Trends Mol Med, 2002, 8(6): 257-260.
[5]	Dheda K, Huggett JF, Bustin SA, et al. Validation of housekeeping genes for normalizing RNA expression in real-time PCR[J]. Biotechniques, 2004, 37(1): 112-119.
[6]	Chi XY, Hu RB, Yang QL, et al. Validation of reference genes for gene expression studies in peanut by quantitative real-time RT-PCR[J]. Mol Genet Genomics, 2012, 287(2): 167-176.
[7]	Lu SF, Sun YH, Shi R, et al. Novel and mechanical stress-responsive microRNAs in Populus trichocarpa that are absent from Arabidopsis[J]. The Plant Cell Online, 2005, 17(8): 2186-2203.
[8]	Zanca AS, Vicentini R, Ortiz-Morea FA, et al. Identification and expression analysis of microRNAs and targets in the biofuel crop sugarcane[J]. BMC Plant Biol, 2010, 10(1): 260.
[9]	Thellin O, Zorzi W, Lakaye B, et al. Housekeeping genes as internal standards: use and limits[J]. J Biotechnol, 1999, 75(2): 291-295.
[10]	Jain M, Nijhawan A, Tyagi AK, et al. Validation of housekeeping genes as internal control for studying gene expression in rice by quantitative real-time PCR[J]. Biochem Biophys Res Commun, 2006, 345(2): 646-651.
[11]	韦朝领, 李叶云, 江昌俊. 茶树逆境生理及其分子生物学研究进展[J]. 安徽农业大学学报, 2009, 36(3): 335-339.
[12]	王新超, 杨亚军. 茶树抗性育种研究现状[J]. 茶叶科学, 2003, 23(2): 94-98.
[13]	李钱峰, 蒋美艳, 于恒秀, 等. 水稻胚乳RNA定量RT-PCR分析中参照基因选择[J]. 扬州大学学报: 农业与生命科学版, 2008, 29(2): 61-66.
[14]	Peltier HJ, Latham GJ.Normalization of microRNA expression levels in quantitative RT-PCR assays: identification of suitable reference RNA targets in normal and cancerous human solid tissues[J]. Rna, 2008, 14(5): 844-852.
[15]	Szabo A, Perou CM, Karaca M, ,et al. Statistical modeling for selecting housekeeper genes [J]. Genome Biol. Statistical modeling for selecting housekeeper genes [J]. Genome Biol, 2004, 5(8): R59.1-R59.10.
[16]	Spanakis E.Problems related to the interpretation of autoradiographic data on gene expression using common constitutive transcripts as controls[J]. Nucleic Acids Res, 1993, 21(16): 3809-3819.
[17]	付媛媛, 穆春生, 高洪文, 等. 紫花苜蓿18 S rRNA基因的克隆及内参基因表达稳定性评价[J]. 植物生理学报, 2014, 50(12): 1809-1815.
[18]	樊连梅, 王超, 刘更森, 等. 苹果着色期实时定量PCR内参基因的筛选和验证[J]. 植物生理学报, 2014, 50(12): 1903-1911.
[19]	严海东, 蒋晓梅, 张新全, 等. 非生物胁迫下多年生黑麦草qRT-PCR分析中内参基因的选择[J]. 农业生物技术学报, 2014, 22(12): 1494-1501.
[20]	Hao XY, Horvath DP, Chao WS, et al. Identification and evaluation of reliable reference genes for quantitative real-time PCR analysis in tea plant (Camellia sinensis (L.) O. Kuntze)[J]. Int J Mol Sci, 2014, 15(12): 22155-22172.
[21]	Monzo M, Navarro A, Bandres E, et al. Overlapping expression of microRNAs in human embryonic colon and colorectal cancer[J]. Cell Res, 2008, 18(8): 823-833.
[22]	Schaefer A, Jung M, Miller K, et al. Suitable reference genes for relative quantification of miRNA expression in prostate cancer[J]. Exp Mol Med, 2010, 42(11): 749-758.
[23]	Wotschofsky Z, Meyer HA, Jung M, et al. Reference genes for the relative quantification of microRNAs in renal cell carcinomas and their metastases[J]. Anal Biochem, 2011, 417(2): 233-241.
[24]	Vandesompele J, Preter KD, Pattyn F, ,et al. Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes [J]. Genome Biol. Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes [J]. Genome Biol, 2002, 3(7): research0034.1-0034.11.

Screening of microRNA Reference Genes for Real-time Fluorescence Quantitative PCR under Cold Stress in Camellia sinensis

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