遗传 ›› 2011, Vol. 33 ›› Issue (11): 1191-1202.doi: 10.3724/SP.J.1005.2011.01191
祁云霞1,2, 刘永斌2, 荣威恒2
收稿日期:2011-01-17
修回日期:2011-04-22
出版日期:2011-11-20
发布日期:2011-11-25
通讯作者:
刘永斌
E-mail:ybliu117@126.com
基金资助:内蒙古自然科学基金项目(编号：2010BS0405)和国家现代肉羊产业技术体系(编号：nycytx-39)资助
QI Yun-Xia1, 2, LIU Yong-Bin2, RONG Wei-Heng2
Received:2011-01-17
Revised:2011-04-22
Online:2011-11-20
Published:2011-11-25
Contact:
LIU Yong-Bin
E-mail:ybliu117@126.com
摘要： 转录组是特定组织或细胞在某一发育阶段或功能状态下转录出来的所有RNA的集合。转录组研究能够从整体水平研究基因功能以及基因结构, 揭示特定生物学过程以及疾病发生过程中的分子机理。RNA-Seq作为一种新的高效、快捷的转录组研究手段正在改变着人们对转录组的认识。RNA-Seq利用高通量测序技术对组织或细胞中所有RNA反转录而成的cDNA文库进行测序, 通过统计相关读段(reads)数计算出不同RNA的表达量, 发现新的转录本; 如果有基因组参考序列, 可以把转录本映射回基因组, 确定转录本位置、剪切情况等更为全面的遗传信息, 已广泛应用于生物学研究、医学研究、临床研究和药物研发等。文章主要介绍了RNA-Seq原理、技术特点及其应用, 并就RNA-Seq技术面临的挑战和未来发展前景进行了讨论, 为今后该技术的研究与应用提供参考。
祁云霞，刘永斌，荣威恒. 转录组研究新技术：RNA-Seq及其应用[J]. 遗传, 2011, 33(11): 1191-1202.
QI Yun-Xia, LIU Yong-Bin, RONG Wei-Heng. RNA-Seq and its applications: a new technology for transcriptomics[J]. HEREDITAS, 2011, 33(11): 1191-1202.
| [1] Lockhart DJ, Winzeler EA. Genomics, gene expression and DNA arrays. Nature, 2000, 405(6788): 827-836.[2] Costa V, Angelini C, De Feis I, Ciccodicola A. Uncovering the complexity of transcriptomes with RNA-Seq. J Biomed Biotechnol, 2010, 2010: 853916.[3] Wang Z, Gerstein M, Snyder M. RNA-Seq: a revolution-ary tool for transcriptomics. Nat Rev Genet, 2009, 10(1): 57-63.[4] 454 Home Page. http://www.454.com/indecx.asp.[5] Illumina Home Page. http://www.illumina.com/.[6] Applied Biosystems Home Page. http://www.appliedbiosystems. com.cn/.[7] Helicos Home Page. http://www.helicosbio.com/.[8] Magi A, Benelli M, Gozzini A, Girolami F, Torricelli F, Brandi ML. Bioinformatics for next generation sequencing data. Genes, 2010, 1(2): 294-307.[9] Nowrousian M. Next-generation sequencing techniques for eukaryotic microorganisms: sequencing-based solutions to biological problems. Eukaryot Cell, 2010, 9(9): 1300-1310.[10] Shendure J, Ji H. Next-generation DNA sequencing. Nat Biotechnol, 2008, 26(10): 1135-1145.[11] Metzker ML. Sequencing technologies-the next generation. Nat Rev Genet, 2010, 11(1): 31-46.[12] Bentley DR, Balasubramanian S, Swerdlow HP, Smith GP, Milton J, Brown CG, Hall KP, Evers DJ, Barnes CL, Bignell HR, Boutell JM, Bryant J, Carter RJ, Keira Cheetham R, Cox AJ. Accurate whole human genome sequencing using reversible terminator chemistry. Nature, 2008, 456(7218): 53-59.[13] Mardis ER. The impact of next-generation sequencing technology on genetics. Trends Genet, 2008, 24(3): 133- 141.[14] Margulies M, Egholm M, Altman WE, Attiya S, Bader JS, Bemben LA, Berka J, Braverman MS, Chen YJ, Chen ZT, Dewell SB, Du L, Fierro JM. Genome sequencing in mi-crofabricated high-density picolitre reactors. Nature, 2005, 437(7057): 376-380.[15] Ronaghi M, Uhlén M, Nyrén P. A sequencing method based on real-time pyrophosphate. Science, 1998, 281(5375): 363-365.[16] Smih DR, Quinlan AR, Peckhham HE, Makowsky K, Tao W, Woolf B, Shen L, Donahue WF, Tusneem N, Stromberg MP, Stewart DA, Zhang L, Ranade SS, Warner JB, Lee CC, Coleman BE, Zhang Z, McLaughlin SF, Malek JA, Sorenson JM, Blanchard AP, Chapman J, Hillman D, Chen F, Rokhsar DS, McKernan KJ, Jeffries TW, Marth GT, Richardson PM. Rapid whole-genome mutational profiling using next-generation sequencing technologies. Genome Res, 2008, 18(10): 1638-1642.[17] Harris TD, Buzby PR, Babcock H, Beer E, Bowers J, Bra-slavsky I, Causey M, Colonell J, Dimeo J, Efcavitch JW, Giladi E, Gill J, Healy J, Jarosz M, Lapen D, Moulton K, Quake SR, Steinmann K, Thayer E, Tyurina A, Ward R, Weiss H, Xie Z. Single-molecule DNA sequencing of a viral genome. Science, 2008, 320(5872): 106-109.[18] Harris TD, Buzby PR, Jarosz M, Gill J, Weiss H, Lapidus SN. Optical train and method for TIRF single molecule detection and analysis. US patent application, 20070070349, 2007.[19] Haas BJ, Zody MC. Advancing RNA-Seq analysis. Nat Biotechnol, 2010, 28(5): 421-423.[20] Ozsolak F, Milos PM. RNA sequencing: advances, chal-lenges and opportunities. Nat Rev Genet, 2011, 12(2): 87-98.[21] Maher CA, Palanisamy N, Brenner JC, Cao XH, Kalyana-Sundaram S, Luo SJ, Khrebtukova I, Barrette TR, Grasso C, Yu JD, Lonigro RJ, Schroth G, Kumar-Sinha C, Chinnaiyan AM. Chimeric transcript discovery by paired-end transcriptome sequencing. Proc Natl Acad Sci USA, 2009, 106(30): 12353-12358.[22] Au KF, Jiang H, Lin L, Xing Y, Wong WH. Detection of splice junctions from paired-end RNA-seq data by SpliceMap. Nucleic Acids Res, 2010, 38(14): 4570-4578.[23] Edgren H, Murumagi A, Kangaspeska S, Nicorici D, Hongisto V, Kleivi K, Rye IH, Nyberg S, Wolf M, Borre-sen-Dale AL, Kallioniemi O. Identification of fusion genes in breast cancer by paired-end RNA-sequencing. Genome Biol, 2011, 12(1): R6.[24] Bashir A, Volik S, Collins C, Bafna V, Raphael BJ. Evaluation of paired-end sequencing strategies for detection of genome rearrangements in cancer. PLoS Comput Biol, 2008, 4(4): e1000051.[25] Okoniewski MJ, Miller CJ. Hybridization interactions between probesets in short oligo microarrays lead to spurious correlations. BMC Bioinformatics, 2006, 7(1): 276.[26] Royce TE, Rozowsky JS, Gerstein MB. Toward a univer-sal microarray: prediction of gene expression through nearest-neighbor probe sequence identification. Nucleic Acids Res, 2007, 35(15): e99.[27] Velculescu VE, Zhang L, Vogelstein B, Kinzler KW. Serial analysis of gene expression. Science, 1995, 270(5235): 484-487.[28] Brenner S, Johnson M, Bridgham J, Golda G, Lloyd DH, Johnson D, Luo SJ, McCurdy S, Foy M, Ewan M, Roth R, George D, Eletr S, Albrecht G, Vermaas E, Williams SR, Moon K, Burcham T, Pallas M, DuBridge RB, Kirchner J, Fearon K, Mao J, Corcoran K. Gene expression analysis by massively parallel signature sequencing (MPSS) on microbead arrays. Nat Biotechnol, 2000, 18(6): 630-634.[29] Wilhelm BT, Marguerat S, Watt S, Schubert F, Wood V, Goodhead I, Penkett CJ, JaneRogers J, Bähler J. Dynamic repertoire of a eukaryotic transcriptome surveyed at single-nucleotide resolution. Nature, 2008, 453(7199): 1239- 1243.[30] Wang XW, Luan JB, Li JM, Bao YY, Zhang CX, Liu SS. De novo characterization of a whitefly transcriptome and analysis of its gene expression during development. BMC Genomics, 2010, 11(1): 400.[31] Xiang LX, He D, Dong WR, Zhang YW, Shao JZ. Deep sequencing-based transcriptome profiling analysis of bacteria-challenged Lateolabrax japonicus reveals insight into the immune-relevant genes in marine fish. BMC Genomics, 2010, 11(1): 472.[32] Vera JC, Wheat CW, Fescemyer HW, Frilander MJ, Crawford DL, Hanski I, Marden JH. Rapid transcriptome characterization for a nonmodel organism using 454 pyrosequencing. Mol Ecol, 2008, 17(7): 1636-1647.[33] Cloonan N, Forrest Alistair RR, Kolle G, Gardiner BBA, Faulkner GJ, Brown MK, Taylor DF, Steptoe AL, Wani S, Bethel G, Robertson AJ, Perkins AC, Bruce SJ, Lee CC, Ranade SS, Peckham HE, Manning JM, McKernan KJ, Grimmond SE. Stem cell transcriptome profiling via massive-scale mRNA sequencing. Nat Methods, 2008, 5(7): 613-619.[34] Morin RD, Bainbridge M, Fejes A, Hirst M, Krzywinski M, Pugh TJ, McDonald H, Varhol R, Jones SJM, Marra MA. Profiling the HeLa S3 transcriptome using randomly primed cDNA and massively parallel short-read sequenc-ing. Biotechniques, 2008, 45(1): 81-94.[35] Nagalakshmi U, Wang Z, Waern K, Shou C, Raha D, Gerstein M, Snyder M. The transcriptional landscape of the yeast genome defined by RNA sequencing. Science, 2008, 320(5881): 1344-1349.[36] Mortazavi A, Williams BA, McCue K, Schaeffer L, Wold B. Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat Methods, 2008, 5(7): 621-628.[37] Zhang GJ, Guo GW, Hu XD, Zhang Y, Li QY, Li RQ, Zhuang RH, Lu ZK, He ZQ, Fang XD, Chen L, Tian W, Tao Y, Kristiansen K, Zhang XQ, Li SG, Yang HM, Wang J, Wang J. Deep RNA sequencing at single base-pair resolution reveals high complexity of the rice transcriptome. Genome Res, 2010, 20(5): 646-654.[38] Lu TT, Lu GJ, Fan DL, Zhu CR, Li W, Zhao Q, Feng Q, Zhao Y, Guo YL, Li WJ, Huang XH, Han B. Function an-notation of the rice transcriptome at single-nucleotide resolution by RNA-seq. Genome Res, 2010, 20(9): 1238-1249.[39] Sultan M, Schulz MH, Richard H, Magen A, Klingenhoff A, Scherf M, Seifert M, Borodina T, Soldatov A, Park-homchuk D, Schmidt D, O'Keeffe S, Haas S, Vingron M, Lehrach H, Yaspo ML. A global view of gene activity and alternative splicing by deep sequencing of the human transcriptome. Science, 2008, 321(5891): 956-960.[40] Campbell MA, Haas BJ, Hamilton JP, Mount SM, Buell CR. Comprehensive analysis of alternative splicing in rice and comparative analyses with Arabidopsis. BMC Genomics, 2006, 7: 327.[41] Wang BB, Brendel V. Genomewide comparative analysis of alternative splicing in plants. Proc Natl Acad Sci USA, 2006, 103(18): 7175-7180.[42] Filichkin SA, Priest HD, Givan SA, Shen R, Bryant DW, Fox SE, Wong WK, Mockler TC. Genome-wide mapping of alternative splicing in Arabidopsis thaliana. Genome Res, 2010, 20(1): 45-58.[43] Chen FC, Wang SS, Chaw SM, Huang YT, Chuang TJ. Plant Gene and Alternatively Spliced Variant Annotator. A plant genome annotation pipeline for rice gene and alter-natively spliced variant identification with cross-species expressed sequence tag conservation from seven plant species. Plant Physiol, 2007, 143(3): 1086-1095.[44] Barbazuk WB, Fu Y, McGinnis KM. Genome-wide analyses of alternative splicing in plants: opportunities and challenges. Genome Res, 2008, 18(9): 1381-1392.[45] Shah SP, Morin RD, Khattra J, Prentice L, Pugh T, Burleigh A, Delaney A, Gelmon K, Guliany R, Senz J, Steidl C, Holt RA, Jones S, Sun M, Leung G, Moore R, Severson T, Taylor GA, Teschendorff AE, Tse K, Turashvili G, Var-hol R, Warren RL, Watson P, Zhao YJ, Caldas C, Hunts-man D, Hirst M, Marra MA, Aparicio S. Mutational evolution in a lobular breast tumour profiled at single nucleo-tide resolution. Nature, 2009, 461(7265): 809-813.[46] Sugarbaker DJ, Richards WG, Gordon GJ, Dong LS, De Rienzo A, Maulik G, Glickman JN, Chirieac LR, Hartman ML, Taillon BE, Du L, Bouffard P, Kingsmore SF, Miller NA, Farmer AD, Jensen RV, Gullans SR, Bueno R. Tran-scriptome sequencing of malignant pleural mesothelioma tumors. Proc Natl Acad Sci USA, 2008, 105(9): 3521-3526.[47] Chepelev I, Wei G, Tang QS, Zhao KJ. Detection of single nucleotide variations in expressed exons of the human genome using RNA-Seq. Nucleic Acids Res, 2009, 37(16): e106.[48] Marioni JC, Mason CE, Mane SM, Stephens M, Gilad Y. RNA-seq: an assessment of technical reproducibility and comparison with gene expression arrays. Genome Res, 2008, 18(9): 1509-1517.[49] Birney E, Stamatoyannopoulos JA, Dutta A, Guigó R, Gingeras TR, Margulies EH, Weng Z, Snyder M, Dermitzakis ET, Thurman RE, Kuehn MS, Taylor CM, Neph S, Koch CM, Asthana S, Malhotra A, Adzhubei I, Greenbaum JA, de Jong PJ. Identification and analysis of functional elements in 1% of the human genome by the ENCODE pilot project. Nature, 2007, 447(7146): 799-816.[50] Clamp M, Fry B, Kamal M, Xie XH, Cuff J, Lin MF, Kellis M, Lindblad-Toh K, Lander ES. Distinguishing pro-tein-coding and noncoding genes in the human genome. Proc Natl Acad Sci USA, 2007, 104(49): 19428-19433.[51] Ponting CP, Oliver PL, Reik W. Evolution and functions of long noncoding RNAs. Cell, 2009, 136(4): 629-641.[52] Filipowicz W, Bhattacharyya SN, Sonenberg N. Mecha-nisms of post-transcriptional regulation by microRNAs: are the answers in sight? Nat Rev Genet, 2008, 9(2): 102-114.[53] Lu C, Tej SS, Luo SJ, Haudenschild CD, Meyers BC, Green PJ. Elucidation of the small RNA component of the transcriptome. Science, 2005, 309(5740): 1567-1569.[54] Xie ZX, Johansen LK, Gustafson AM, Kasschau KD, Lel-lis AD, Zilberman D, Jacobsen SE, Carrington JC. Genetic and functional diversication of small RNA pathways in plants. PloS Biol, 2004, 2(5): 642-652.[55] Zhao T, Li GL, Mi SJ, Li S, Hannon GJ, Wang XJ, Qi YJ. A complex system of small RNAs in the unicellular green alga Chlamydomonas reinhardtii. Genes Dev, 2007, 21(10): 1190-1203.[56] Burnside J, Bernberg E, Anderson A, Lu C, Meyers BC, Green PJ, Jain N, Isaacs G, Morgan RW. Marek's disease virus encodes microRNAs that map to meq and the latency-associated transcript. J Virol, 2006, 80(17): 8778-8786.[57] Yao YY, Guo GG, Ni ZF, Sunkar R, Du JK, Zhu JK, Sun QX. Cloning and characterization of microRNAs from wheat (Triticum aestivum L.). Genome Biol, 2007, 8(6): R96.[58] Berezikov E, Thuemmler F, van Laake LW, Kondova I, Bontrop R, Cuppen E, Plasterk RH. Diversity of microRNAs in human and chimpanzee brain. Nat Genet, 2006, 38(12): 1375-1377.[59] Lau NC, Seto AG, Kim J, Kuramochi-Miyagawa S, Na-kano T, Bartel DP, Kingston RE. Characterization of the piRNA complex from rat testes. Science, 2006, 313(5785): 363-367.[60] Morin RD, O'Connor MD, Griffith M, Kuchenbauer F, Delaney A, Prabhu AL, Zhao YJ, McDonald H, Zeng T, Hirst M, Eaves CJ, Marra MA. Application of massively parallel sequencing to microRNA profiling and discovery in human embryonic stem cells. Genome Res, 2008, 18(4): 610-621.[61] Guttman M, Amit I, Garber M, French C, Lin MF, Feldser D, Huarte M, Zuk O, Carey BW, Cassady JP, Cabili MN, Jaenisch R, Mikkelsen TS, Jacks T, Hacohen N, Bernstein BE, Kellis M, Regev A, Rinn JL, Lander ES. Chromatin signature reveals over a thousand highly conserved large non-coding RNAs in mammals. Nature, 2009, 458(7235): 223-227.[62] Khalil AM, Guttman M, Huarte M, Garber M, Raj A, Rivea Morales D, Thomas K, Presser A, Bernstein BE, van Oudenaarden A, Regev A, Lander ES, Rinn JL. Many human large intergenic noncoding RNAs associate with chromatin-modifying complexes and affect gene expression. Proc Natl Acad Sci USA, 2009, 106(28): 11667-11672.[63] 黄文涛, 郭向前, 戴甲培, 陈润生. MicroRNA, lncRNA与神经退行性疾病. 生物化学与生物物理进展, 2010, 37(8): 826-833.[64] Wilusz JE, Sunwoo H, Spector DL. Long noncoding RNAs: functional surprises from the RNA world. Genes Dev, 2009, 23(13): 1494-1504.[65] Peng XX, Gralinski L, Armour CD, Ferris MT, Thomas MJ, Proll S, Bradel-Tretheway BG, Korth MJ, Castle JC, Biery MC, Bouzek HK, Haynor DR, Frieman MB, Heise M, Raymond CK, Baric RS, Katze MG. Unique signatures of long noncoding RNA expression in response to virus infection and altered innate immune signaling. MBio, 2010, 1(5): e00206-10.[66] Lister R, O'Malley RC, Tonti-Filippini J, Gregory BD, Berry CC, Millar AH, Ecker JR. Highly integrated single-base resolution maps of the epigenome in Arabidopsis. Cell, 2008, 133(3): 523-536.[67] Bruno VM, Wang Z, Marjani SL, Euskirchen GM, Martin J, Sherlock G, Snyder M. Comprehensive annotation of the transcriptome of the human fungal pathogen Candida albicans using RNA-seq. Genome Res, 2010, 20(10): 1451-1458.[68] Vliet VA. Next generation sequencing of microbial tran-scriptomes: challenges and opportunities. FEMS Microbiol Lett, 2010, 302(1): 1-7.[69] Tang FC, Barbacioru C, Wang YZ, Nordman E, Lee C, Xu NL, Wang XH, Bodeau J, Tuch BB, Siddiqui A, Lao KQ, Surani MA. mRNA-Seq whole-transcriptome analysis of a single cell. Nat Methods, 2009, 6(5): 377-382.[70] Croucher NJ, Fookes MC, Perkins TT, Turner DJ, Mar-guerat SB, Keane T, Quail MA, He M, Assefa S, Bähler J, Kingsley RA, Parkhill J, Bentley SD, Dougan G, Thomson NR. A simple method for directional transcriptome sequencing using Illumina technology. Nucleic Acids Res, 2009, 37(22): e148.[71] Vivancos AP, Güell M, Dohm JC, Serrano L, Himmel-bauer H. Strand-specific deep sequencing of the tran-scriptome. Genome Res, 2010, 20(7): 989-999. |
| [1] | 石田培,张莉. 全转录组学在畜牧业中的应用[J]. 遗传, 2019, 41(3): 193-205. |
| [2] | 魏金川,徐添翼,吴静,宋晓峰. 真核生物基因组长内含子递归剪接事件的分子机制[J]. 遗传, 2019, 41(2): 89-97. |
| [3] | 张高华, 于树涛, 王鹤, 王旭达. 高油酸花生发芽期低温胁迫转录组及差异表达基因分析[J]. 遗传, 2019, 41(11): 1050-1059. |
| [4] | 张宇,白素英,马跃. 麝鼠前列腺在繁殖期和非繁殖期转录组差异分析[J]. 遗传, 2018, 40(6): 488-495. |
| [5] | 任岚,肖茹丹,张倩,娄晓敏,张昭军,方向东. KLF1和KLF9对K562细胞红系分化的协同调控作用[J]. 遗传, 2018, 40(11): 998-1006. |
| [6] | 杨莹,陈宇晟,孙宝发,杨运桂. RNA甲基化修饰调控和规律[J]. 遗传, 2018, 40(11): 964-976. |
| [7] | 刘亚军,张峰,刘宏德,孙啸. 下一代测序技术在干细胞转录调控研究中的应用[J]. 遗传, 2017, 39(8): 717-725. |
| [8] | 黄万龙,张秀秀,李嫒,苗向阳. 利用RNA-seq技术筛选大白猪皮下和肌内脂肪 组织差异表达基因[J]. 遗传, 2017, 39(6): 501-511. |
| [9] | 叶仲杰,刘启鹏,岑山,李晓宇. LINE-1编码的逆转录酶在肿瘤形成过程中的作用[J]. 遗传, 2017, 39(5): 368-376. |
| [10] | 魏凯,马磊. 高通量测序时代下持家基因定义的发展[J]. 遗传, 2017, 39(2): 127-134. |
| [11] | 李光奇, 孙从佼, 吴桂琴, 石凤英, 刘爱巧, 孙皓, 杨宁. 利用转录组测序筛选鸡蛋褐壳性状相关基因[J]. 遗传, 2017, 39(11): 1102-1111. |
| [12] | 刘永明, 张玲, 邱涛, 赵卓凡, 曹墨菊. 高通量转录组测序技术在植物雄性不育研究中的应用[J]. 遗传, 2016, 38(8): 677-687. |
| [13] | 朱帅旗, 龚一富, 杭雨晴, 刘浩, 王何瑜. 绿色杜氏藻转录组分析[J]. 遗传, 2015, 37(8): 828-836. |
| [14] | 王冬, 李永君, 丁楠, 王均云, 杨琼, 杨雅冉, 李艳明, 方向东, 赵华. miRNA调控恶性黑色素瘤细胞上皮-间充质转化的分子网络及机制[J]. 遗传, 2015, 37(7): 673-682. |
| [15] | 谢兵兵, 杨亚东, 丁楠, 方向东. 整合分析多组学数据筛选疾病靶点的精准医学策略[J]. 遗传, 2015, 37(7): 655-663. |
| 阅读次数 | ||||||
|
全文 |
|
|||||
|
摘要 |
|
|||||
www.chinagene.cn
备案号：京ICP备09063187号-4
总访问:,今日访问:,当前在线:
