葵百合LsGRP1及其同源基因之研究

LsGRP1 and its homologues in Lilium cv. Star Gazer

吳詩敏(Shih-Min Wu)

指導教授 ： 陳昭瑩

國立臺灣大學/生物資源暨農學院/植物病理與微生物學研究所/碩士(2006年)

https://doi.org/10.6342/NTU.2006.02097

摘要

葵百合LsGRP1（Lilium cv. Star Gazer glycine-rich protein 1）cDNA，為利用抗病誘導物質水楊酸（salicylic acid，SA），以PCR-select subtraction法及differential screening篩選得到的防禦相關基因，並已推測LsGRP1在百合由水楊酸與病原菌誘導的防禦反應中為重要的因子。本研究利用北方雜合分析偵測葵百合接受不同刺激之LsGRP1的表現量，結果顯示除了水楊酸，甲基茉莉酸和胺基環丙烷羧酸也會誘導LsGRP1的表現。為得到LsGRP1訊息RNA的全長度cDNA，利用快速擴增cDNA末端法，得到LsGRP1之5’端未轉譯區，包含起始密碼前21 bp；並得到相似性序列，推測葵百合有LsGRP1同源基因。分別在葵百合基因組DNA及訊息RNA上找尋LsGRP1同源基因，於前者進行聚合酵素連鎖反應，結果發現葵百合LsGRP1基因中包含了180 bp的隱子（intron），此外並發現利用不同引子對，以基因組DNA與訊息RNA分別進行基因組與反轉錄聚合酵素連鎖反應均得到多重產物；其中得到一cDNA較LsGRP1多出208 bp的重複性序列，命名為LsGRP2。另一以3’端快速擴增cDNA末端法得到之同源基因LsGRP3，僅可轉譯出70個胺基酸。由5’端與3’快速擴增cDNA法得到比LsGRP1多39 bp重複序列的LsGRP4，其轉譯出的胺基酸序列與LsGRP1相似度為90%。本研究指出葵百合LsGRP1於基因體中存在同源基因，然此等基因是否表現或其表現情形並不清楚。

關鍵字

葵百合 ； glycine-rich-protein ； LsGRP1 ； 同源基因

並列摘要

A cDNA, named LsGRP1, has been obtained by suppression subtractive hybridization from salicylic acid (SA)-treated Lilum cv. Star Gazer plants, followed by differential screening. In the previous study, LsGRP1 was presumed to play a role in SA and pathogen-induced defense responses in lily. In this study, the expression of LsGRP1 in ‘Star Gazer’ leaves was analyzed by Northern blot analysis. The result showed that not only SA, but also methyl jasmonate and 1-aminocyclopropane-1-
carboxylic acid can induce LsGRP1 expression. Rapid amplification of cDNA ends was performed to obtain 5’-untranslated region of LsGRP1,that is 21-bp upstream of the start codon. In addition, the homologue of LsGRP1 was investigated on the genome and mRNA of ‘Star Gazer’. By the way, LsGRP1 gene was amplified from genomic DNA of ‘Star Gazer’ by polymerase chain reaction (PCR), which contains a 180-bp intron. On the other hand, multiple fragments were amplified from genomic DNA and mRNA by PCR and reverse transcription-PCR using different primer pairs. A fragment which a 208-bp repeated sequence of LsGRP1 was named LsGRP2. The second homologues of LsGRP1, named LsGRP3, only encoded 70 amino acid residues. The third homologue was 39-bp longer than LsGRP1, encoding a peptide sequence of 90% identity to that of LsGRP1. These results indicated that homologues of LsGRP1 present in the genome of Lilium cv. Star Gazer although the expression of these homologues has not yet be analyzed.

並列關鍵字

Lilium cv. Star Gazer ； glycine-rich protein ； LsGRP1 ； homologues

參考文獻

Aneeta, Sanan-Mishra, N., Tuteja, N., and Kumar Sopory, S. 2002. Salinity- and ABA-induced up-regulation and light-mediated modulation of mRNA encoding glycine-rich RNA-binding protein from Sorghum bicolor. Biochem. Biophys. Res. Commun. 296:1063-1068.

Carpenter, C. D., Kreps, J. A., and Simon, A. E. 1994. Genes encoding glycine-rich Arabidopsis thaliana proteins with RNA-binding motifs are influenced by cold treatment and an endogenous circadian rhythm. Plant Physiol. 104:1015-1025.

Cassab, G. I. 1998. Plant cell wall proteins. Annu. Rev. Plant Physiol. Plant Mol. Biol. 49:281-309.

Condit, C. M., and Meagher, R. B. 1986. A gene encoding a novel glycine-rich structural protein of petunia. Nature 323:178-181.

Condit, C. M., G., M. B., and B., M. R. 1990. Characterization of the expression of the petunia glycine-rich protein-1 gene product. Plant Physiol. 93:596-602.