SSH法获取水稻矮化突变体相关的cDNA片段

Identification of cDNA Fragments Differentially Expressed Between a Rice Mutant dwarf69 and Wild Type TGA-2 by Suppressive Subtraction Hybridization

利用抑制性消减杂交 (Suppressivesubtractionhybridization ,SSH)分离水稻矮化突变体dwarf69与其野生型对照品系特光矮 2 (Oryzasativa ,TGA 2indica)间表达有差异的cDNA片段。分别以dwarf69作为驱赶子 ,TGA 2为检测子 ,以及以dwarf69作为检测子 ,TGA 2为驱赶子建立了两个差异表达cDNA文库 ,分别代表在TGA 2和dwarf69特异表达的cDNA。经反式Northern检测这两个文库共得到 1 0个阳性克隆。序列分析表明有两个序列为水稻中首次报道 ,一个序列有相应的基因组序列报道 ,但其cDNA序列是首次报道。

The suppressive subtraction hybridization (SSH) technique was used to isolate the cDNA fragments that showed differential expression between the rice mutant dwarf69 and wild type strain,TGA 2(( Oryza sativa, indica).Two differential expressing cDNA libraries were constructed using the cDNAs of dwarf69 as driver and the cDNAs of TGA 2 as tester, vice versa. These two cDNA libraries indicated the differential expressing cDNA fragments in TGA 2 and dwarf69. By reverse Northern blots, ten positive clones were obtained. Subsequently sequencing analysis showed two cDNA fragments were first reported in rice, and another was reported as genomic sequence previously but not as cDNA.