綠竹中 BoMSP41 基因之分子選殖與檢定

Molecular cloning and characterization of BoMSP41 in Bambusa oldhamii

張皓鈞(Hao-Chun Chang)

指導教授 ： 王愛玉

國立臺灣大學/生命科學院/生化科技學系/碩士(2012年)

https://doi.org/10.6342/NTU.2012.10871

摘要

竹類植物通常具有快速生長的特性。本研究以綠竹 (Bambusa oldhamii) 的cDNA以及genomic DNA為模版進行PCR，選殖出一個在快速延長的綠竹節間受到正調控的未知功能的ORF。將其cDNA及genomic DNA序列進行比對，可以發現兩者之核苷酸序列及胺基酸序列不完全相同但同質度高，顯示兩者為同源基因，並且得知在ORF中沒有intron的存在；兩者的序列轉譯後得到的蛋白質分子量皆為40.8 kDa ，序列長度為354個胺基酸。以BLAST進行分析發現，在目前的基因庫中只比對到單子葉植物的相似序列，並且功能未知，因此將此二同源基因分別命名為BoMSP41-1 (Bambusa oldhamii中的蛋白質monocot specific protein，分子量為41 kDa) 及BoMSP41-2。兩種BoMSP41胺基酸序列中皆富含Gln、Glu以及其他帶電荷的胺基酸Lys、Arg、Asp，但是缺少Trp。此外，亦存在一些重複序列。藉由PONDR軟體預測結果，顯示兩種BoMSP41都具有一段的結構不穩定區段，因此推測其為intrinsically disordered proteins。另外BoMSP41-1的表現以real-time PCR分析，發現BoMSP41-1mRNA在快速延長中的節間中的表現量大幅增加，因此推測該基因在綠竹生長中扮演了重要的角色。

關鍵字

竹生長發育 ； 綠竹 ； 單子葉植物 ； 快速生長 ； 無穩定構型蛋白質

並列摘要

Growth of bamboo is generally faster than other plants. The coding region of an function-unknown gene, which was shown to be up-regulated in the rapidly elongating bamboo culms, was cloned from the shoots of Bambusa oldhamii using PCR with genomic DNA and cDNAs from bamboo shoots as templates. Alignment of the genomic sequence and cDNA sequence of this gene showed that they were homologs with high identities in both the nucleotide and amino acid sequences, and there was no intron within the open reading frames. Both sequences encoded a 40.8-kDa polypeptide of 354 amino acid residues. BLAST searching revealed that they might be monocot-specific and that the function of there homologs in other plants remained unknown. They were designated BoMSP41-1 (for monocot-specific protein of 41 kDa in Bambusa oldhamii) and BoMSP41-2, respectively. The deduced amino acid sequences of BoMSP41-1 and BoMSP41-2 were rich in Gln and Glu, and other charged amino acid residues Lys, Arg and Asp, but was lacking inTrp. In addition, several tandemly repeated sequences were identify . A long disorder region with high probability was predicted in each BoMSP41 by the disorder prediction program PONDR, suggesting that they were intrinsically disordered proteins. The expression of BoMSP41-2 analyzed by real-time RT-PCR showed that the increase in BoMSP41-1 mRNA abundance paralleled the elongation rate of bamboo culms, indicating that the gene plays an important role in bamboo growth.

並列關鍵字

bamboo growth ； monocot ； Bambusa MSP41 ； Bambusa oldhamii ； intrinsically disordered protein

參考文獻

Brown CJ, Johnson AK, Dunker AK, Daughdrill GW (2011) Evolution and disorder. Curr Opin Struc Biol 21: 441-446

Burnette WN (1981) "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Analytical Biochemistry 112: 195-203

Dunker AK (2004) Instrinsically disordered protein and cell signaling. Biophysical Journal 86: 354a-355a

Dunker AK, Brown CJ, Lawson JD, Iakoucheva-Sebat LM, Vucetic S, Obradovic Z (2002a) The protein trinity: Structure/function relationships that include intrinsic disorder. Biophysical Journal 82: 460a-460a

Dunker AK, Brown CJ, Lawson JD, Iakoucheva LM, Obradovic Z (2002b) Intrinsic disorder and protein function. Biochemistry 41: 6573-6582