Please use this identifier to cite or link to this item:
標題: 1.大量表現miR160a與miR167b對水稻生長發育影響之探討2.miRNA對其目標基因調控快速檢測方法的建立
1. Effects on rice growth and development by overexpressing miR160a and miR167b 2. A quick method for detecting miRNA target genes
作者: 林宣妤
Lin, Syuan-Yu
關鍵字: rice;水稻;microRNA;OsARF;microRNA;OsARF
出版社: 分子生物學研究所
引用: 鄭至忠 (2008). 大量表現miR160a與miR167b對水稻生長發育之影響. 國立中興大學分子生物學研究所 碩士學位論文. Abdel-Ghany, S.E., and Pilon, M. (2008). MicroRNA-mediated systemic down-regulation of copper protein expression in response to low copper availability in Arabidopsis. J Biol Chem 283, 15932-15945. Abel, S., and Theologis, A. (1996). Early genes and auxin action. Plant Physiol 111, 9-17. Achard, M.E., Tree, J.J., Holden, J.A., Simpfendorfer, K.R., Wijburg, O.L., Strugnell, R.A., Schembri, M.A., Sweet, M.J., Jennings, M.P., and McEwan, A.G. (2010). The multi-copper-ion oxidase CueO of Salmonella enterica serovar Typhimurium is required for systemic virulence. Infect Immun 78, 2312-2319. Adenot, X., Elmayan, T., Lauressergues, D., Boutet, S., Bouche, N., Gasciolli, V., and Vaucheret, H. (2006). DRB4-dependent TAS3 trans-acting siRNAs control leaf morphology through AGO7. Curr Biol 16, 927-932. Archak, S., and Nagaraju, J. (2007). Computational prediction of rice (Oryza sativa) miRNA targets. Genomics Proteomics Bioinformatics 5, 196-206. Baumberger, N., and Baulcombe, D.C. (2005). Arabidopsis ARGONAUTE1 is an RNA Slicer that selectively recruits microRNAs and short interfering RNAs. Proc Natl Acad Sci U S A 102, 11928-11933. Bozhkov, P.V., Suarez, M.F., Filonova, L.H., Daniel, G., Zamyatnin, A.A., Jr., Rodriguez-Nieto, S., Zhivotovsky, B., and Smertenko, A. (2005). Cysteine protease mcII-Pa executes programmed cell death during plant embryogenesis. Proc Natl Acad Sci U S A 102, 14463-14468. Brady, S.M., Sarkar, S.F., Bonetta, D., and McCourt, P. (2003). The ABSCISIC ACID INSENSITIVE 3 (ABI3) gene is modulated by farnesylation and is involved in auxin signaling and lateral root development in Arabidopsis. Plant J 34, 67-75. Capron, A., Gourgues, M., Neiva, L.S., Faure, J.E., Berger, F., Pagnussat, G., Krishnan, A., Alvarez-Mejia, C., Vielle-Calzada, J.P., Lee, Y.R., et al. (2008). Maternal control of male-gamete delivery in Arabidopsis involves a putative GPI-anchored protein encoded by the LORELEI gene. Plant Cell 20, 3038-3049. Chen, X. (2004). A microRNA as a translational repressor of APETALA2 in Arabidopsis flower development. Science 303, 2022-2025. Chen, X. (2009). Small RNAs and their roles in plant development. Annu Rev Cell Dev Biol 25, 21-44. Crocoll, C., Asbach, J., Novak, J., Gershenzon, J., and Degenhardt, J. (2010). Terpene synthases of oregano (Origanum vulgare L.) and their roles in the pathway and regulation of terpene biosynthesis. Plant Mol Biol 73, 587-603. Davies, P.J. (1995). Plant hormones: Physiology, biochemistry and molecular biology. Kluwer, London, UK ed. 2. Dharmasiri, N., Dharmasiri, S., Jones, A.M., and Estelle, M. (2003). Auxin action in a cell-free system. Curr Biol 13, 1418-1422. Gandikota, M., Birkenbihl, R.P., Hohmann, S., Cardon, G.H., Saedler, H., and Huijser, P. (2007). The miRNA156/157 recognition element in the 3'' UTR of the Arabidopsis SBP box gene SPL3 prevents early flowering by translational inhibition in seedlings. Plant J 49, 683-693. Garcia, D., Collier, S.A., Byrne, M.E., and Martienssen, R.A. (2006). Specification of leaf polarity in Arabidopsis via the trans-acting siRNA pathway. Curr Biol 16, 933-938. Garg, R., Jhanwar, S., Tyagi, A.K., and Jain, M. (2010). Genome-wide survey and expression analysis suggest diverse roles of glutaredoxin gene family members during development and response to various stimuli in rice. DNA Res 17, 353-367. Gifford, M.L., Dean, A., Gutierrez, R.A., Coruzzi, G.M., and Birnbaum, K.D. (2008). Cell-specific nitrogen responses mediate developmental plasticity. Proc Natl Acad Sci U S A 105, 803-808. Guilfoyle, T.J., and Hagen, G. (2001 ). Auxin response factors. J Plant Growth Reg 10, 281-291. Gutierrez, L., Bussell, J.D., Pacurar, D.I., Schwambach, J., Pacurar, M., and Bellini, C. (2009). Phenotypic plasticity of adventitious rooting in Arabidopsis is controlled by complex regulation of AUXIN RESPONSE FACTOR transcripts and microRNA abundance. Plant Cell 21, 3119-3132. Hardtke, C.S., and Berleth, T. (1998). The Arabidopsis gene MONOPTEROS encodes a transcription factor mediating embryo axis formation and vascular development. EMBO J 17, 1405-1411. Jacobsen, S.E., Running, M.P., and Meyerowitz, E.M. (1999). Disruption of an RNA helicase/RNAse III gene in Arabidopsis causes unregulated cell division in floral meristems. Development 126, 5231-5243. Jin, J., Huang, W., Gao, J.P., Yang, J., Shi, M., Zhu, M.Z., Luo, D., and Lin, H.X. (2008). Genetic control of rice plant architecture under domestication. Nat Genet 40, 1365-1369. Jones-Rhoades, M.W., and Bartel, D.P. (2004). Computational identification of plant microRNAs and their targets, including a stress-induced miRNA. Mol Cell 14, 787-799. Jones-Rhoades, M.W., Bartel, D.P., and Bartel, B. (2006). MicroRNAS and their regulatory roles in plants. Annu Rev Plant Biol 57, 19-53. Jung, H.J., and Kang, H. (2007). Expression and functional analyses of microRNA417 in Arabidopsis thaliana under stress conditions. Plant Physiol Biochem 45, 805-811. Kepinski, S., and Leyser, O. (2004). Auxin-induced SCFTIR1-Aux/IAA interaction involves stable modification of the SCFTIR1 complex. Proc Natl Acad Sci U S A 101, 12381-12386. Khraiwesh, B., Arif, M.A., Seumel, G.I., Ossowski, S., Weigel, D., Reski, R., and Frank, W. (2010). Transcriptional control of gene expression by microRNAs. Cell 140, 111-122. Kim, M.C., Lee, S.H., Kim, J.K., Chun, H.J., Choi, M.S., Chung, W.S., Moon, B.C., Kang, C.H., Park, C.Y., Yoo, J.H., et al. (2002). Mlo, a modulator of plant defense and cell death, is a novel calmodulin-binding protein. Isolation and characterization of a rice Mlo homologue. J Biol Chem 277, 19304-19314. Lee, K.O., Lee, J.R., Yoo, J.Y., Jang, H.H., Moon, J.C., Jung, B.G., Chi, Y.H., Park, S.K., Lee, S.S., Lim, C.O., et al. (2002). GSH-dependent peroxidase activity of the rice (Oryza sativa) glutaredoxin, a thioltransferase. Biochem Biophys Res Commun 296, 1152-1156. Lee, R.C., Feinbaum, R.L., and Ambros, V. (1993). The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell 75, 843-854. Lee, Y., Bak, G., Choi, Y., Chuang, W.I., and Cho, H.T. (2008). Roles of phosphatidylinositol 3-kinase in root hair growth. Plant Physiol 147, 624-635. Leshem, Y., Seri, L., and Levine, A. (2007). Induction of phosphatidylinositol 3-kinase-mediated endocytosis by salt stress leads to intracellular production of reactive oxygen species and salt tolerance. Plant J 51, 185-197. Li, D., Liu, H., Zhang, H., Wang, X., and Song, F. (2008a). OsBIRH1, a DEAD-box RNA helicase with functions in modulating defence responses against pathogen infection and oxidative stress. J Exp Bot 59, 2133-2146. Li, J., Dai, X., and Zhao, Y. (2006). A role for auxin response factor 19 in auxin and ethylene signaling in Arabidopsis. Plant Physiol 140, 899-908. Li, P., Wang, Y., Qian, Q., Fu, Z., Wang, M., Zeng, D., Li, B., Wang, X., and Li, J. (2007). LAZY1 controls rice shoot gravitropism through regulating polar auxin transport. Cell Res 17, 402-410. Li, T., Li, H., Zhang, Y.X., and Liu, J.Y. (2010a). Identification and analysis of seven H2O2-responsive miRNAs and 32 new miRNAs in the seedlings of rice (Oryza sativa L. ssp. indica). Nucleic Acids Res. Li, W.X., Oono, Y., Zhu, J., He, X.J., Wu, J.M., Iida, K., Lu, X.Y., Cui, X., Jin, H., and Zhu, J.K. (2008b). The Arabidopsis NFYA5 transcription factor is regulated transcriptionally and posttranscriptionally to promote drought resistance. Plant Cell 20, 2238-2251. Li, Y.F., Zheng, Y., Addo-Quaye, C., Zhang, L., Saini, A., Jagadeeswaran, G., Axtell, M.J., Zhang, W., and Sunkar, R. (2010b). Transcriptome-wide identification of microRNA targets in rice. Plant J 62, 742-759. Liu, P.P., Montgomery, T.A., Fahlgren, N., Kasschau, K.D., Nonogaki, H., and Carrington, J.C. (2007). Repression of AUXIN RESPONSE FACTOR10 by microRNA160 is critical for seed germination and post-germination stages. Plant J 52, 133-146. Liu, Q., and Chen, Y.Q. (2009). Insights into the mechanism of plant development: interactions of miRNAs pathway with phytohormone response. Biochem Biophys Res Commun 384, 1-5. Lobbestael, E., Reumers, V., Ibrahimi, A., Paesen, K., Thiry, I., Gijsbers, R., Van den Haute, C., Debyser, Z., Baekelandt, V., and Taymans, J.M. (2010). Immunohistochemical detection of transgene expression in the brain using small epitope tags. BMC Biotechnol 10, 16. Luo, Y.C., Zhou, H., Li, Y., Chen, J.Y., Yang, J.H., Chen, Y.Q., and Qu, L.H. (2006). Rice embryogenic calli express a unique set of microRNAs, suggesting regulatory roles of microRNAs in plant post-embryogenic development. FEBS Lett 580, 5111-5116. Mallory, A.C., Bartel, D.P., and Bartel, B. (2005). MicroRNA-directed regulation of Arabidopsis AUXIN RESPONSE FACTOR17 is essential for proper development and modulates expression of early auxin response genes. Plant Cell 17, 1360-1375. Meng, Y., Huang, F., Shi, Q., Cao, J., Chen, D., Zhang, J., Ni, J., Wu, P., and Chen, M. (2009). Genome-wide survey of rice microRNAs and microRNA-target pairs in the root of a novel auxin-resistant mutant. Planta 230, 883-898. Nag, R., Maity, M.K., and Dasgupta, M. (2005). Dual DNA binding property of ABA insensitive 3 like factors targeted to promoters responsive to ABA and auxin. Plant Mol Biol 59, 821-838. Nagpal, P., Ellis, C.M., Weber, H., Ploense, S.E., Barkawi, L.S., Guilfoyle, T.J., Hagen, G., Alonso, J.M., Cohen, J.D., Farmer, E.E., et al. (2005). Auxin response factors ARF6 and ARF8 promote jasmonic acid production and flower maturation. Development 132, 4107-4118. Nakamura, T., and Sugita, M. (2008). A conserved DYW domain of the pentatricopeptide repeat protein possesses a novel endoribonuclease activity. FEBS Lett 582, 4163-4168. Nodine, M.D., and Bartel, D.P. (2010). MicroRNAs prevent precocious gene expression and enable pattern formation during plant embryogenesis. Genes Dev 24, 2678-2692. Ohnishi, T., Yokota, T., and Mizutani, M. (2009). Insights into the function and evolution of P450s in plant steroid metabolism. Phytochemistry 70, 1918-1929. Okushima, Y., Overvoorde, P.J., Arima, K., Alonso, J.M., Chan, A., Chang, C., Ecker, J.R., Hughes, B., Lui, A., Nguyen, D., et al. (2005). Functional genomic analysis of the AUXIN RESPONSE FACTOR gene family members in Arabidopsis thaliana: unique and overlapping functions of ARF7 and ARF19. Plant Cell 17, 444-463. Potuschak, T., Vansiri, A., Binder, B.M., Lechner, E., Vierstra, R.D., and Genschik, P. (2006). The exoribonuclease XRN4 is a component of the ethylene response pathway in Arabidopsis. Plant Cell 18, 3047-3057. Preger, V., Tango, N., Marchand, C., Lemaire, S.D., Carbonera, D., Di Valentin, M., Costa, A., Pupillo, P., and Trost, P. (2009). Auxin-responsive genes AIR12 code for a new family of plasma membrane b-type cytochromes specific to flowering plants. Plant Physiol 150, 606-620. Qi, Y., Denli, A.M., and Hannon, G.J. (2005). Biochemical specialization within Arabidopsis RNA silencing pathways. Mol Cell 19, 421-428. Radwan, O., Mouzeyar, S., Nicolas, P., and Bouzidi, M.F. (2005). Induction of a sunflower CC-NBS-LRR resistance gene analogue during incompatible interaction with Plasmopara halstedii. J Exp Bot 56, 567-575. Raghavendra, A.S., Gonugunta, V.K., Christmann, A., and Grill, E. (2010). ABA perception and signalling. Trends Plant Sci 15, 395-401. Reinhart, B.J., Weinstein, E.G., Rhoades, M.W., Bartel, B., and Bartel, D.P. (2002). MicroRNAs in plants. Genes Dev 16, 1616-1626. Reyes, J.L., and Chua, N.H. (2007). ABA induction of miR159 controls transcript levels of two MYB factors during Arabidopsis seed germination. Plant J 49, 592-606. Schneider, K., Mathur, J., Boudonck, K., Wells, B., Dolan, L., and Roberts, K. (1998). The ROOT HAIRLESS 1 gene encodes a nuclear protein required for root hair initiation in Arabidopsis. Genes Dev 12, 2013-2021. Schwarz, S., Grande, A.V., Bujdoso, N., Saedler, H., and Huijser, P. (2008). The microRNA regulated SBP-box genes SPL9 and SPL15 control shoot maturation in Arabidopsis. Plant Mol Biol 67, 183-195. Serpeloni, M., Vidal, N.M., Goldenberg, S., Avila, A.R., and Hoffmann, F.G. (2011). Comparative genomics of proteins involved in RNA nucleocytoplasmic export. BMC Evol Biol 11, 7. Shen, J., Xie, K., and Xiong, L. (2010). Global expression profiling of rice microRNAs by one-tube stem-loop reverse transcription quantitative PCR revealed important roles of microRNAs in abiotic stress responses. Mol Genet Genomics 284, 477-488. Singh, U., Deb, D., Singh, A., and Grover, A. (2011). Glycine-rich RNA binding protein of Oryza sativa inhibits growth of M15 E. coli cells. BMC Res Notes 4, 18. Skowyra, D., Craig, K.L., Tyers, M., Elledge, S.J., and Harper, J.W. (1997). F-box proteins are receptors that recruit phosphorylated substrates to the SCF ubiquitin-ligase complex. Cell 91, 209-219. Southern, J.A., Young, D.F., Heaney, F., Baumgartner, W.K., and Randall, R.E. (1991). Identification of an epitope on the P and V proteins of simian virus 5 that distinguishes between two isolates with different biological characteristics. J Gen Virol 72 ( Pt 7), 1551-1557. Sozzani, R., Maggio, C., Varotto, S., Canova, S., Bergounioux, C., Albani, D., and Cella, R. (2006). Interplay between Arabidopsis activating factors E2Fb and E2Fa in cell cycle progression and development. Plant Physiol 140, 1355-1366. Sunkar, R., Girke, T., Jain, P.K., and Zhu, J.K. (2005). Cloning and characterization of microRNAs from rice. Plant Cell 17, 1397-1411. Sunkar, R., Zhou, X., Zheng, Y., Zhang, W., and Zhu, J.K. (2008). Identification of novel and candidate miRNAs in rice by high throughput sequencing. BMC Plant Biol 8, 25. Tian, C.E., Muto, H., Higuchi, K., Matamura, T., Tatematsu, K., Koshiba, T., and Yamamoto, K.T. (2004). Disruption and overexpression of auxin response factor 8 gene of Arabidopsis affect hypocotyl elongation and root growth habit, indicating its possible involvement in auxin homeostasis in light condition. Plant J 40, 333-343. Tiwari, S.B., Hagen, G., and Guilfoyle, T. (2003). The roles of auxin response factor domains in auxin-responsive transcription. Plant Cell 15, 533-543. Ulmasov, T., Hagen, G., and Guilfoyle, T.J. (1997). ARF1, a transcription factor that binds to auxin response elements. Science 276, 1865-1868. Ulmasov, T., Hagen, G., and Guilfoyle, T.J. (1999). Dimerization and DNA binding of auxin response factors. Plant J 19, 309-319. Ulmasov, T., Liu, Z.B., Hagen, G., and Guilfoyle, T.J. (1995). Composite structure of auxin response elements. Plant Cell 7, 1611-1623. Waller, F., Furuya, M., and Nick, P. (2002). OsARF1, an auxin response factor from rice, is auxin-regulated and classifies as a primary auxin responsive gene. Plant Mol Biol 50, 415-425. Wan, X., Mo, A., Liu, S., Yang, L., and Li, L. (2010). Constitutive expression of a peanut ubiquitin-conjugating enzyme gene in Arabidopsis confers improved water-stress tolerance through regulation of stress-responsive gene expression. J Biosci Bioeng. Wang, D., Pei, K., Fu, Y., Sun, Z., Li, S., Liu, H., Tang, K., Han, B., and Tao, Y. (2007). Genome-wide analysis of the auxin response factors (ARF) gene family in rice (Oryza sativa). Gene 394, 13-24. Wang, J.W., Wang, L.J., Mao, Y.B., Cai, W.J., Xue, H.W., and Chen, X.Y. (2005). Control of root cap formation by MicroRNA-targeted auxin response factors in Arabidopsis. Plant Cell 17, 2204-2216. Wang, L., Gu, X., Xu, D., Wang, W., Wang, H., Zeng, M., Chang, Z., Huang, H., and Cui, X. (2011). miR396-targeted AtGRF transcription factors are required for coordination of cell division and differentiation during leaf development in Arabidopsis. J Exp Bot 62, 761-773. Wang, L., Xu, Y., Zhang, C., Ma, Q., Joo, S.H., Kim, S.K., Xu, Z., and Chong, K. (2008). OsLIC, a Novel CCCH-Type Zinc Finger Protein with Transcription Activation, Mediates Rice Architecture via Brassinosteroids Signaling. PLoS One 3, e3521. Wang, X.J., Reyes, J.L., Chua, N.H., and Gaasterland, T. (2004). Prediction and identification of Arabidopsis thaliana microRNAs and their mRNA targets. Genome Biol 5, R65. Weijers, D., and Jurgens, G. (2004). Funneling auxin action: specificity in signal transduction. Curr Opin Plant Biol 7, 687-693. Wenzel, C.L., Schuetz, M., Yu, Q., and Mattsson, J. (2007). Dynamics of MONOPTEROS and PIN-FORMED1 expression during leaf vein pattern formation in Arabidopsis thaliana. Plant J 49, 387-398. Wilmoth, J.C., Wang, S., Tiwari, S.B., Joshi, A.D., Hagen, G., Guilfoyle, T.J., Alonso, J.M., Ecker, J.R., and Reed, J.W. (2005). NPH4/ARF7 and ARF19 promote leaf expansion and auxin-induced lateral root formation. Plant J 43, 118-130. Wu, L., Zhang, Q., Zhou, H., Ni, F., Wu, X., and Qi, Y. (2009). Rice MicroRNA effector complexes and targets. Plant Cell 21, 3421-3435. Wu, M.F., Tian, Q., and Reed, J.W. (2006). Arabidopsis microRNA167 controls patterns of ARF6 and ARF8 expression, and regulates both female and male reproduction. Development 133, 4211-4218. Xie, K., Wu, C., and Xiong, L. (2006). Genomic organization, differential expression, and interaction of SQUAMOSA promoter-binding-like transcription factors and microRNA156 in rice. Plant Physiol 142, 280-293. Xie, Z., Khanna, K., and Ruan, S. (2010). Expression of microRNAs and its regulation in plants. Semin Cell Dev Biol 21, 790-797. Yan, Y.S., Chen, X.Y., Yang, K., Sun, Z.X., Fu, Y.P., Zhang, Y.M., and Fang, R.X. (2010). Overexpression of an F-box Protein Gene Reduces Abiotic Stress Tolerance and Promotes Root Growth in Rice. Mol Plant. Yang, J.H., Han, S.J., Yoon, E.K., and Lee, W.S. (2006). Evidence of an auxin signal pathway, microRNA167-ARF8-GH3, and its response to exogenous auxin in cultured rice cells. Nucleic Acids Res 34, 1892-1899. Yoshihara, T., and Iino, M. (2007). Identification of the gravitropism-related rice gene LAZY1 and elucidation of LAZY1-dependent and -independent gravity signaling pathways. Plant Cell Physiol 48, 678-688. Yu, B., Lin, Z., Li, H., Li, X., Li, J., Wang, Y., Zhang, X., Zhu, Z., Zhai, W., Wang, X., et al. (2007). TAC1, a major quantitative trait locus controlling tiller angle in rice. Plant J 52, 891-898. Zhao, B., Ge, L., Liang, R., Li, W., Ruan, K., Lin, H., and Jin, Y. (2009). Members of miR-169 family are induced by high salinity and transiently inhibit the NF-YA transcription factor. BMC Mol Biol 10, 29. Zhao, B., Liang, R., Ge, L., Li, W., Xiao, H., Lin, H., Ruan, K., and Jin, Y. (2007). Identification of drought-induced microRNAs in rice. Biochem Biophys Res Commun 354, 585-590. Zhou, L., Liu, Y., Liu, Z., Kong, D., Duan, M., and Luo, L. (2010). Genome-wide identification and analysis of drought-responsive microRNAs in Oryza sativa. J Exp Bot 61, 4157-4168. Zhou, W., Dong, L., Ginsburg, D., Bouhassira, E.E., and Tsai, H.M. (2005). Enzymatically active ADAMTS13 variants are not inhibited by anti-ADAMTS13 autoantibodies: a novel therapeutic strategy? J Biol Chem 280, 39934-39941. Zhu, Q.H., Spriggs, A., Matthew, L., Fan, L., Kennedy, G., Gubler, F., and Helliwell, C. (2008). A diverse set of microRNAs and microRNA-like small RNAs in developing rice grains. Genome Res 18, 1456-1465.
MicroRNA(miRNA)是一群大小約21nt的單股RNA,在生物體中會藉著與目標基因mRNA配對而對目標基因mRNA進行剪切,或是抑制轉譯作用來達到調控基因表現的目的。目前已知miRNA在植物的生長發育扮演著重要的角色,包括藉著抑制某些轉錄因子來調控下游基因的表現,Auxin response factor(ARF)就是其中一群轉錄因子。水稻中已經有許多miRNA被發現,但只有少數被深入研究。本研究室過去發現大量表現miR160a和miR167b的水稻轉殖株(35S::miR160a和35S::miR167b)會影響目標基因ARF之表現,以RLM-5’RACE證實miR160a會對目標基因OsARF18與OsARF22 mRNA進行剪切而抑制其表現量。本研究觀察到在ABA的處理下,轉殖株35S::miR160a幼苗根部生長被抑制的情形較Wild-type嚴重,顯示大量表現miR160a可能增加了植株幼苗根部對ABA的敏感度。另外還觀察到從孕穗期開始,轉殖株的分蘗角度較大,顯示大量表現miR160a可能改變了植株的株型。miR167b經前人以軟體比對發現其目標基因可能為ARF25,但在轉植株35S::miR167b中兩者並無明顯消長的情形。本研究檢測轉植株35S::miR167b五個獨立株系後代的miR167b及ARF25表現量,結果顯示miR167b表現量越高的轉殖株,其ARF25表現量越低,可見ARF25很有可能是miR167b的目標基因。對轉殖株進行性狀觀察,發現其中三個miR167b表現量為Wild-type 4~7倍的品系,性狀與Wild-type相較之下無明顯差異,但另外兩個miR167b表現量為Wild-type 9~11倍的品系,則有分蘗數較少且分蘗角度較大的性狀,顯示miR167b表現量的增加或ARF25表現量降低可能與此性狀有關。另外,本研究以miR160a對OsARF18的調節作用為依據,擬建構miRNA調節其目標基因表現的檢測方法,但目前尚未獲得明確及穩定之結果。

MiRNA (miRNAs), a group of single-stranded small RNAs, are about 21 nucleotides in length. These miRNAs regulate their target genes by mRNA cleavage or transcriptional repression. MiRNAs play an important role in plant growth and development, including down-regulating transcription factors to operate the transcript levels of downstream genes. Auxin response factors (ARFs) are one of the transcription factor families which are regulated by miRNAs. There have been many miRNAs found in rice, but few of them have been carefully investigated. It has been shown that transgenic rice over-expressing miR160a could suppress the expressions of OsARF18 and OsARF22 genes, and their targeted-cleavage sites has been identified by RLM-5'RACE assay. Further treatment with exogenously added ABA showed more severe inhibition on seedling roots growth in miR160a over-expression transgenic rice than those of the wild-type. In addition, a wider tiller growth angle since the booting stage was observed in miR160a over-expression transgenic rice suggesting a regulatory role of miR160a in rice architecture development. Another rice miR167b has been studied to analyze its effects on plant growth and target gene OsARF25 by creating miR167b over-expression transgenic rice. The expression levels of miR167b and its putative target gene OsARF25 of five transgenic rice lines were analyzed. Results showed that the higher the expression levels of miR167b in the transgenic rice lines, the lower the expression levels of OsARF25 were detected, suggesting that OsARF25 is the target gene of miR167b. A phenotype with dwarf and wider tiller growth angle was observed in two independent transgenic lines that expressed approximately 9 to 11-fold of miR167b than that of the wild-type, while the other three lines that expressed approximately 4 to 7-fold of miR167b revealed the same phenotype as the wild-type. How can this phenotype be attributed by the different expression levels of miR167b and OsARF25 require further investigation. In this study, an approach to establish a quick miRNA target genes screening method using the known relationship between miR160a and OsARF18 was also performed, but was not successful. Further investigations to increase the sensitivity and stability of the assay method are required.
其他識別: U0005-1002201112181100
Appears in Collections:分子生物學研究所

Show full item record

Google ScholarTM


Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.