野生稻OrufipogonW19431888条全长cDNA序列的数据分析.ppt

野生稻O. rufipogon W1943 1888 条全长cDNA序列的数据分析 NCGR 2008-09-03 背景 野生稻O. rufipogon（AA genome）是与栽 培稻关系最近的祖先水稻品种1,2。 具有许多优于栽培稻的农艺性状，比如耐旱、 耐盐等等3,4； 公共数据库中有大量栽培稻的基因组序列信息 5,6，同时也有大量的cDNA资源7,8； 极少野生稻的序列和克隆资源，比较成规模的 是Oryza minuta (BBCC genome) 5,211条 叶片ests9。 现状与目的 NCGR野生稻资源：克隆并精确测序了1,888个 unique的O. rufipogon W1943 cDNA克隆。 期望通过W1943 cDNA序列与籼、粳稻cDNA 序列的比较： 汇总一些水稻新基因、潜在野生稻特有的基因、 W1943特有剪切方式基因、组织特异性高表达 的基因和与microRNA相关的基因； 提供一些线索，供有兴趣者作进一步研究之用。 1888 W1943 cDNAs BLAST against cultivated rice genomic sequences and cDNAs 1888 W1943 cDNAs SSR comparison with indica and japonica cDNAs 一、未匹配粳稻基因组之基因 定义：未能定位到O. sativa japonica Nipponbare genome sequences，但与籼稻93-11基因 组序列有同源或与水稻ests序列有同源或与其它禾本科ests序列有同源。 且去除与细菌有同源的基 因 解释：或者落于粳稻基因组测序gap中，或者籼稻特有的基因，或者野生稻特有基因。 name93-11 contigsESTs or mRNA hits proteinname93-11 contigsESTs or mRNA hits protein CT842002Contig005912AK241925-CU406895Contig003011CT859459- CT842007Contig008507CT856206-CU861744Contig000750AK099287ring-box protein CU405940Contig001402AK103326-CU405657-CT856885- CU406172Contig014596AK242967-CT841712-CA766528- CT842006Contig000383AK111647GTP-binding protein CU405768-CT83665660S ribosomal protein L7A CU861753Contig000750AK099287ring-box protein CU405675-CA75623560S ribosomal protein L17 CU406308Contig000444AK070131-CU406202-NM_001063334- CT841996Contig002576CT834800-CU406924-AC145809- CU406568Contig003848AK064050Bowman Birk trypsin inhibitor CU405898-CN130755.1 (Sorghum bicolor) ribulose- bisphosphate carboxylase CU406582Contig000444AK107776-CU406778-BE429292.1 (Triticum turgidum) - CU406596Contig001277AK242711-CU861677-FF534517.1 (Manihot esculenta) - CT842008Contig008507CT856206-CT841912-EH277383.1 (Spartina alterniflora) - 二、水稻新基因 定义：能定位到栽培稻基因组序列的同源，但无任何已知水稻表达序列的同 源。与rice MPSS搜索比较几乎没有找到匹配片段。 解释：水稻新基因。或者在栽培稻中表达量过低难于克隆，或者野生稻特有 。 nameLen(bp)Chr location Identity(%)nameLen(bp)Chr location Identity(%)Antisenseprotein CU4069106561099CU4057857270599CA764081DNA-directed RNA polymerase 3 CU4061385680299CU8617954750979CT858901- CU4060225431299CU4063558371297AK107125AP2 domain, putative CU40575747704100CU4063965200299AK103485- CU40692141402100CT8418009411199AK121962patatin, putative CU40653538902100CU8616886930899AK109182- CU4068325301092CT84193715520898AK106713- CU4068714580184 注：该17个基因均没有找到任何蛋白同源匹配。右侧的7个基因与已知 的水稻表达序列成反义RNA对。 CU8618043830699 CU86172155401100 三、W1943特有剪切方式基因 定义：与栽培稻japonica基因组序列完全一致(100% identity)，同时与栽培稻表达序 列同源但剪切方式独有(独特的AS剪切方式)。 解释：或只是尚未克隆到该AS表达方式；或为野生稻所独具。 nameLen(bp)Chr locationNo. of exonprotein CT841942978076 (1st intron: GC-AG)- CU406810958066 (1st intron: GT-TG)dual-specificity phosphatase protein CT8418931011016drought-induced protein CT8418741369014vesicle transport protein CU4058531377051dehydration-responsive protein CU405923639071IAA amidohydrolase CU406279648051- CU406025839021- CT841561740062- CU406579468092- CU4069351345012- CU4066001107012- CU405570952012- CU406091893013- CU406134665103- some W1943 cDNAs unique splicing pattern : The expression level of every gene should exceed 100 tpm (times per million) of at least one tissue. : If the gene expressed in several diverse tissues, then the percentage of the highest expression level should be more than 75% among all tissues. : The ratio of the first two highest expression level should be over 10. 41 putative tissue-specific genes10 /rice/ exon剪切出现intron；intron中出现exon exon剪切出现intron intergenic区出现exon 2个exon合并成单exon 四、组织特异性高表达基因 namelenORFtissueprotein orw1943s101k15619bp51-434bpleaflight-regulated protein orw1943c102c24833bp62-463bpleafsubunit of ribulose-1,5-bisphosphate carboxylase orw1943s101p081544bp111-650bpleafglycolate oxidase orw1943s101h18912bp120-752bpleafH+-transporting ATP synthase chain 9-like protein orw1943c113b17837bp108-623bpleafretrotransposon protein, putative, unclassified orw1943c002g13843bp58-576bpleafalanine aminotransferase orw1943c003d24404bp18-227bpleafferredoxin-NADP(H) oxidoreductase orw1943c104a05985bp70-777bpleafphotosystem-1 F subunit precursor orw1943c006o21625bp89-310bprootmetallothionein-like protein type 1 orw1943c103g16916bp110-553bprootMAPEG family orw1943c003o09619bo111-311bprootPotato inhibitor I family orw1943c112g141008bp58-825bprootreceptor-like protein kinase orw1943c104g04805bp115-618bprootpathogenesis-related protein PR-1a orw1943c006h22664bp1-453bprootpathogenesis-related protein 4b orw1943s101p06512bp36-353bprootN-carbamyl-L-amino acid amidohydrolase orw1943c111d191399bp59-1312bpgerminating seedlingputative alpha-galactosidase orw1943c002o05769bp82-432bpgerminating seedlingnonspecific lipid-transfer protein 2 precursor orw1943c002p22518bp92-331bpmeristematic tissuemetallothionein orw1943c109d02682bp45-230bpmuture pollenputative lipase 五、潜在miRNA及miRNA靶基因 microRNAs：21-23nt 小分子RNA，由具发夹 结构的70-90nt单链 RNA前体经Dicer酶加工 而来。具种间保守性。 作用方式：通过不完全 互补结合到靶目标 mRNA (多数3 UTR区) ，诱发蛋白翻译抑制， 不影响转录本的稳定性 ；少数miRNA可能以类 似siRNA的方式诱导靶 目标mRNA的降解。根 据互补的完全程度发挥 不同的作用。 判断流程及标准11： 鉴定潜在的野生稻miRNA putative miRNALength(bp)pre-miRNA len(bp)hit_miRNAChr locationmiRNA sequence CU4062921416262osa-MIR159a01uuuggauugaagggagcucug CU4059431511101osa-MIR156j06ugacagaagagagugagcac CU86181956180osa-miR818e04aaucccuuauauuuugggacgg CU861752727150osa-miR44610aucaauaugaaugugggaaau CU406292 pre-miRNA CU405943 pre-miRNA reference 1.Wang, Z. Y., Second, G., and Tanksley, S. D. 1992, Polymorphism and phylogenetic relationships among species in the genus Oryza as determined by analysis of nuclear RFLPs, Theor. Appl. Genet., 83, 565581. 2.Londo, J. P., Chiang, Y. C., Hung, K. H., Chiang, T. Y., and Schaal, B. A. 2006, Phylogeography of Asian wild rice, Oryza rufipogon, reveals multiple independent domestications of cultivated rice, Oryza sativa, Proc. Natl. Acad. Sci. USA, 103, 9578-83. 3.Zhang, X., Zhou, S., Fu, Y., Su, Z., Wang, X., and Sun, C. 2006, Identification of a drought tolerant introgression line derived from Dongxiang common wild rice (O. rufipogon Griff.), Plant Mol. Biol., 62, 247-59. 4.Tian, F., Zhu, Z., Zhang, B., et al. 2006, Fine mapping of a quantitative trait locus for grain number per panicle from wild rice (Oryza rufipogon Griff.), Theor. Appl. Genet., 113, 619-29. 5.International Rice Genome Sequencing Project. (2005), The map-based sequence of the rice genome, Nature, 436, 793-800. 6.Yu, J., Hu, S., Wang, J., et al. 2002, A draft sequence of the rice genome Oryza sativa L.ssp. indica, Science, 296, 92-100. 7.The Rice Full-Length cDNA Consortium. (2003), Collection, mapping, and annotation of over 28,000 cDNA clones from japonica rice, Science, 301, 376-379. 8.Liu, X. H., Lu, T. T., Yu, S. L., et al. 2007, A collection of 10,096 indica rice f