胎兔皮肤组织蛋白质组双向电泳技术的建立与分析  

Establishment and analysis of two-dimensional electrophoresis for proteomes of fetal rabbit skins

作　　者：邱俊[1] 周继红[1] 张波[1] 

机构地区：[1]解放军第三军医大学野战外科研究所四室

出　　处：《中国临床康复》2005年第14期56-57,i002,共3页Chinese Journal of Clinical Rehabilitation

基　　金：国家"九七三"项目资助(G1999054200)~~

摘　　要：目的:胎兔皮肤蛋白质组的研究可能解开其无瘢痕愈合的机制,建立重复性好、分辨率高的胎兔皮肤组织蛋白质组双向电泳技术,初步分析寻找皮肤切割伤伤后胎兔蛋白质表达的差异。方法:本实验于2004-01/12在第三军医大学野战外科研究所完成。取孕20d的新西兰大耳白兔、行宫内手术致胎兔皮肤全层切割伤,24h后取致伤胎兔及其同胞未致伤胎兔背部皮肤,分别确定为致伤组和未致伤组,应用哺乳动物总蛋白提取试剂盒提取其总蛋白。应用双向电泳法将致伤组和未致伤组分别制成3块固相pH梯度胶条,电泳结束后硝酸银染色、图像扫描。同一胶条进行3次双向电泳,对所得图像进行蛋白质点、匹配率以及伤后蛋白表达差异的分析。结果:致伤组和未致伤组胎兔皮肤组织总蛋白分别分离出蛋白质点(105±14)个、(108±19)个,在两组中各选1块胶作为参考胶进行3块胶间的匹配,平均匹配点数分别是(88±8)个、(90±7)个,匹配率分别为84%、83%,同一样本3块胶上不同蛋白质点在第一向的位置偏差分别为(0.85±0.25)mm、(0.9±0.4)mm,在第二向的偏差分别为(1.25±0.5)mm、(1.5±0.5)mm。通过比对得到胎兔伤后表达增加的蛋白质点20个。结论:加大皮肤组织总蛋白上样量进行双向电泳得到重复性和分辨率均较好的电泳图谱。AIM:To establish technique of two dimensional electrophoresis in good repetit ion and high resolution for proteomic analysis of fetal rabbit skins and primari ly analyze and investigate the difference of protein expression in incised wound of fetal rabbit skins based on that the study on proteomes of rabbit skins can reveal the mechanism of scarless healing. METHODS:The experiment was performed in the Research Institute of Surgery,Thir d Military Medical University of Chinese PLA from January to December 2004.Intra uterine operation was conducted to New Zealand flap eared white rabbits pregnan t for 20 days to cause incised wound of full thickness skins of fetal rabbits.A fter 24 hours,skins of back were sampled from the wounded rabbits and those woun dless rabbits as wounded group and woundless group respectively.The complete mam malian proteome extraction kit was used to extract the proteome of fetal rabbit skins.Three pieces of immobilized pH gradient strips were made in the two groups respectively by two dimensiond electrophoresis technique.Two dimensiond elect rophoresis was followed by silver nitrate staining and image scanning.Two dimen siond electrophoresis was used to any piece of mmobized pH gradrent strips for t hree times to analyze the proteome spots,match rate of the gotten images and the expression difference of post wound proteomes. RESULTS:It was detected with this technique that the protein was (105±14) and (108±19) spots separated from the proteomes of fetal rabbit skins in wounded g roup and woundless group respectively.One piece of immobilized pH gradient strip from both groups were selected as the control to the two others,and the mean ma tch spots were 88±8 and 90±7 in the rate of 84%and 83%respectively.The prote in deviation distance of spots in the 3 pieces of mmobized pH gradrent strips fr om the same skin was (0.85±0.25) mm,(0.9±0.4) mm for the first dimensional ele ctrophoresis,and (1.25±0.5) mm,(1.5±0.5) mm for the second dimensional electro phoresis.Twenty differential protei

关 键 词：创伤 动物 皮肤 蛋白质类/分析 电泳 

分 类 号：R318[医药卫生—生物医学工程]