Screening and Analysis of Proteins Interacting with TaPDK from Physiological Male Sterility Induced by CHA in Wheat  

作　　者：ZHANG Long-yu ZHANG Gai-sheng ZHAO Xin-liang YANG Shu-ling 

机构地区：[1]College of Agronomy, Northwest A&F University/Wheat Breeding Engineering Research Center, Ministry of Education/Key Laboratory of Crop Heterosis of Shaanxi Province

出　　处：《Journal of Integrative Agriculture》2013年第6期941-950,共10页农业科学学报（英文版）

基　　金：supported by the National High-Tech R&D Program of China(2011AA10A106);the National Natural Science Foundation of China(31071477,31171611);the Key Scientific and Technological Innovation Special Projects of Shaanxi"13115",China(2010ZDKG-68,2011KTZB02-01-01)

摘　　要：To further research the regulatory network of pyruvate dehydrogenase kinase （designated as TaPDK） in physiological male-sterility （PHYMS） of wheat induced by chemical hybridizing agent （CHA） SQ-1, an anther cDNA library was constructed, and the proteins interacting with TaPDK were screened via yeast two-hybrid technique. Subsequently, a few candidate proteins in nucleotide expression levels were detected by real-time quantitative PCR. Yeast-two hybrid screening was performed by mating yeast strain Y2HGold containing BD-TaPDK bait plasmid with yeast strain Y187 including anther cDNA library plasmid. Diploid yeast cells were plated on synthetic dropout nutrient medium （SD/-Ade/-His/-Leu/-Trp） （QDO）, and further were incubated on QDO medium containing AbA and X-α-Gal. The interactions between TaPDK and the proteins obtained from positive colonies were further confirmed by co-transformation validation. After plasmids DNA were extracted from blue colonies and sequenced, the sequences results were analyzed by bioinformatic methods. Finally, 24 colonies were obtained, including eight genes, namely non-specific lipid-transfer protein precursor （TanLTP）, polyubiquitin （TaPUbi）, glyceraldehyde-3-phosphate dehydrogenase, proliferating cell nuclear antigen （TaPCNA）, CBS domain containing protein （TaCBS）, actin, guanine nucleotide-binding protein beta subunit, chalcone synthase, and three new genes with unknown function. The results of quantitative RT-PCR showed that the expression levels of TanLTP, TaPUbi, and TaPCNA were obviously up-regulated in PHYMS anther, and TaCBS expression was only increased at the tricellular stage in PHYMS anther compared with in fertile lines. Whereas, the expression of TaPDK was obviously down-regulated in PHYMS lines. Collectively, these datas indicated that the majority of candidate proteins might be related to pollen abortion in PHYMS lines, which further suggested that TaPDK plays multiple roles in pollen development, besides participating in regulating pTo further research the regulatory network of pyruvate dehydrogenase kinase （designated as TaPDK） in physiological male-sterility （PHYMS） of wheat induced by chemical hybridizing agent （CHA） SQ-1, an anther cDNA library was constructed, and the proteins interacting with TaPDK were screened via yeast two-hybrid technique. Subsequently, a few candidate proteins in nucleotide expression levels were detected by real-time quantitative PCR. Yeast-two hybrid screening was performed by mating yeast strain Y2HGold containing BD-TaPDK bait plasmid with yeast strain Y187 including anther cDNA library plasmid. Diploid yeast cells were plated on synthetic dropout nutrient medium （SD/-Ade/-His/-Leu/-Trp） （QDO）, and further were incubated on QDO medium containing AbA and X-α-Gal. The interactions between TaPDK and the proteins obtained from positive colonies were further confirmed by co-transformation validation. After plasmids DNA were extracted from blue colonies and sequenced, the sequences results were analyzed by bioinformatic methods. Finally, 24 colonies were obtained, including eight genes, namely non-specific lipid-transfer protein precursor （TanLTP）, polyubiquitin （TaPUbi）, glyceraldehyde-3-phosphate dehydrogenase, proliferating cell nuclear antigen （TaPCNA）, CBS domain containing protein （TaCBS）, actin, guanine nucleotide-binding protein beta subunit, chalcone synthase, and three new genes with unknown function. The results of quantitative RT-PCR showed that the expression levels of TanLTP, TaPUbi, and TaPCNA were obviously up-regulated in PHYMS anther, and TaCBS expression was only increased at the tricellular stage in PHYMS anther compared with in fertile lines. Whereas, the expression of TaPDK was obviously down-regulated in PHYMS lines. Collectively, these datas indicated that the majority of candidate proteins might be related to pollen abortion in PHYMS lines, which further suggested that TaPDK plays multiple roles in pollen development, besides participating in regulating p

关 键 词：wheat （Triticum aestivum L.） chemical hybridizing agent pyruvate dehydrogenase kinase yeast two-hybrid pollen abortion 

分 类 号：S512.1[农业科学—作物学]