桃果实硬核后期中果皮与内果皮中蛋白质组表达的双向电泳分析

A Two-dimensional Electrophoresis Protocol to study Protein Discrepancies in Mesocarp and Putamen during prunus persica Fruit Development

为了探究桃果实发育过程后期中、内果皮不同的代谢进程,以＂大久保＂桃（Prunus persica L.cv.Okubao）盛花后56天的果实为试材,TCA/丙酮沉淀法提取蛋白质,采用固相pH梯度（immobilized pHgradients,IPG）双向电泳系统分离蛋白质,凝胶银染显色后用图像分析软件比较分析差异表达的蛋白点,初步了解中果皮和内果皮蛋白质组表达的差异点。采用pH5~8IPG胶条进行等电聚焦得到的双向电泳图谱在中果皮与内果皮中检测到蛋白质点的数量分别为（1273±101）和（1292±115）个,所有匹配的蛋白点中有65个在中果皮与内果皮的图谱中均存在,其中30个为内果皮相对于中果皮2倍（P〈0.05）上调的点,35个为内果皮相对于中果皮2倍（P〈0.05）下调的点。有10个蛋白点仅在中果皮表达,18个蛋白点只在内果皮表达而中果皮未显示。表明桃果实发育硬核期中果皮与内果皮的蛋白质组存在一定差异,这些差异可能会为进一步解释中果皮与内果皮在代谢的方向上不同而提供理论依据。

Peach （Prunus persica L.cv.Okubao） fruits around 56 days after anthesis were used in this experiment in order to realize the different development of its mesocarp and endocarp.The total protein was extracted by TCA/ cold acetone precipitation,and isolated by immobilized pH gradient （ IPG ） based on 2-DE routine.After silver staining,the gels were evaluated through operating image analysis software and different protein spots were found in mesocarp and sclerised endocarp.（1273±101） and （1292±115） protein spots were detected respectively from 2-DE patterns of mesocarp and endocarp that pH 5-8 IPG strips were practised in the process of soelectric focusing.65 spots of aggregate matched protein spots was present in both patterns of mesocarp and endocarp,and 30 spots showed 2-fold （ t-test,P0.05 ） up-regulated on quantitative appearing an increase in spot intensity and maximum abundance in sclerised endocarp than mesocarp,in addition 45 protein spots expressed 2-fold （ t-test,P0.05 ） down-regulated.10 spots were just observed in mesocarp ’spattern,whereas 18 spots only found in the pattern of endocarp.The results suggest that the proteome component in mesocarp and sclerised endocarp of peach during fruit development took on some discrepancies,and these diversities may be provide an explanation and foundation data ulteriorly about different direction of metabolic models in mesocarp and endocarp.