杜梨干旱胁迫相关水通道蛋白基因的筛选与克隆

以杜梨实生苗为材料,采用15%PEG 6000进行模拟干旱处理,利用qRTPCR、同源重组、亚细胞定位等方法,研究了不同水通道蛋白基因在15%PEG 6000胁迫条件下表达水平的变化趋势,筛选了响应干旱胁迫相关的水通道蛋白基因,并对其进行克隆和分析,以期为梨抗旱基因筛选提供参考依据。结果表明:AQPs在杜梨的根、茎、叶中均有表达,且随着胁迫时间的延长,大部分AQPs在根系中的表达水平呈先上升后下降的趋势,其中PIP2;2基因在胁迫处理3h后,相对表达量最高;从杜梨根系中克隆获得PbPIP2;2,该基因包含一个完整的846bp的开放阅读框,编码281个氨基酸。氨基酸序列比对分析结果表明,PbPIP2;2与其它近缘物种具有一致的保守功能域,其中与苹果MdPIP2;2、白梨PcPIP2;2相似度最高。洋葱表皮亚细胞定位发现PbPIP2;2基因定位在质膜上。

Screening and Cloning of Drought Stress Related Aquaporin Genes in Pyrusbetulifolia Bunge

The seedlings of Pyrus betulifolia Bunge were used as materials,drought treatment was simulated by adding 15% PEG 6000 in the nutrient solution.The changes of expression levels of different aquaporin genes under 15% PEG6000 stress were studied by qRT-PCR,homologous recombination and subcellular localization methods,in order to provide reference for selection of drought resistant genes in pear.The results showed that multiple aquaporin genes were expressed in root,stem and leaf,and the expression levels of most of the aquaporin genes in root increased and then declined with the prolongation of stress time.Among them,PIP2;2 gene had the highest relative expression level after 3hours stress treatment.Then PIP2;2 gene was cloned from root of Pyrus betulifolia Bunge and the gene contained a complete open reading frame of 846 bp encoding 281 amino acids.The alignment of the amino acid sequence showed that PbPIP2;2 had a highly conserved functional domain similar to other related species,with the highest similarity with MdPIP2;2 and PcPIP2;2.By onion epidermis subcellular positioning analysis,we observed that the protein encoded by PbPIP2;2 was localized on the plasma membrane.