Kollipara Krishna P, Saab Imad N, Wych Robert D, Lauer Michael J, Singletary George W
Pioneer Hi-Bred International, Johnston, Iowa 50131-1004, USA.
Plant Physiol. 2002 Jul;129(3):974-92. doi: 10.1104/pp.000729.
Recombinant inbred lines (RILs) derived from B73 x M017 were screened for cold germination (CG) and desiccation tolerance (DT) phenotypes. Reciprocal F(1) hybrids were made between divergent RILs, and hybrids that showed differential phenotypes (parent-of-origin effect) for CG or DT were selected for profiling mRNA and protein expression. mRNA and proteins were extracted from embryo axes of seed germinated for 11 d at 12.5 degrees C in the dark and developing embryos at 40% seed moisture (R5 stage) for CG and DT, respectively. GeneCalling analysis, an open-ended mRNA profiling method, identified 336 of 32,496 and 656 of 32,940 cDNA fragments that showed >or=1.5-fold change in expression between the reciprocal F(1) hybrids for CG and DT, respectively. Protein expression map (PEM) analysis, an open-ended two-dimensional polyacrylamide gel electrophoresis, identified 117 of 2,641 and 205 of 1,876 detected proteins to be differentially expressed with >or=1.5-fold change between the reciprocal F(1) hybrids in CG and DT samples, respectively. A subset of these proteins was identified by tandem mass spectrometry followed by database query of the spectra. The differentially expressed genes/proteins were classified into various functional groups including carbohydrate and amino acid metabolism, ion transporters, stress and defense response, polyamine metabolism, chaperonins, cytoskeleton associated, etc. Phenotypic analysis of seed from self-pollinated ears of the reciprocal F(1) hybrids displayed small differences compared with the reciprocal hybrids themselves, suggesting a negligible effect of cytoplasmic factors on CG and DT traits. The results provide leads to improving our understanding of the genes involved in stress response during seed maturation and germination.
对由B73×M017衍生的重组自交系(RILs)进行了冷萌发(CG)和耐干燥性(DT)表型筛选。在不同的RILs之间构建了正反交F1杂种,并选择在CG或DT方面表现出不同表型(亲本来源效应)的杂种进行mRNA和蛋白质表达分析。分别从在12.5℃黑暗条件下萌发11天的种子胚轴以及种子含水量为40%(R5阶段)的发育胚中提取用于CG和DT分析的mRNA和蛋白质。基因表达谱分析(GeneCalling analysis)是一种开放式mRNA分析方法,在正反交F1杂种之间,分别在32496个和32940个cDNA片段中鉴定出336个和656个在CG和DT中表达变化≥1.5倍的片段。蛋白质表达图谱(PEM)分析是一种开放式二维聚丙烯酰胺凝胶电泳,在正反交F1杂种之间,分别在检测到的2641个和1876个蛋白质中鉴定出117个和205个表达差异≥1.5倍的蛋白质。通过串联质谱对这些蛋白质的一个子集进行鉴定,随后对光谱进行数据库查询。差异表达的基因/蛋白质被分类到各种功能组中,包括碳水化合物和氨基酸代谢、离子转运蛋白、应激和防御反应、多胺代谢、伴侣蛋白、细胞骨架相关蛋白等。正反交F1杂种自交穗上种子的表型分析显示,与正反交杂种本身相比差异较小,表明细胞质因子对CG和DT性状的影响可忽略不计。这些结果为增进我们对种子成熟和萌发过程中应激反应相关基因的理解提供了线索。