牛血清清蛋白在小鼠骨肉瘤模型PCR基因分型中的应用  

Application of bovine serum albumin in the PCR genotyping of osteosarcoma mouse models

作　　者：谷俊侠[1] 乔龙威[1] 梁玉婷[1] 冉德园[1] 吕耀娟[1,2] 郑其平[1,2] 

机构地区：[1]江苏大学基础医学与医学技术学院,江苏镇江212013 [2]Rush大学医学中心,美国芝加哥60612

出　　处：《临床检验杂志》2013年第11期844-847,共4页Chinese Journal of Clinical Laboratory Science

基　　金：国家自然科学基金(31271399)

摘　　要：目的验证在PCR反应体系中加入牛血清清蛋白(BSA)可否提高小鼠骨肉瘤模型floxed Rb1基因杂合子和纯合子的检出率与准确率。方法以小鼠骨肉瘤模型子代小鼠基因组DNA为模板,进行Rb1基因的PCR基因分型。对在PCR反应体系中添加适量浓度(0.16 mg/mL)的BSA与不加BSA的PCR基因分型结果相比较。PCR引物序列涵盖Rb1基因外显子19两侧或3'端的LoxP序列。结果反应体系中添加BSA后,可成功获得PCR扩增条带:单一650 bp或247 bp(野生型)、单一700bp或295 bp(floxed Rb1基因纯合型)以及650 bp和700 bp(或247 bp和295 bp)混合条带(floxed Rb1基因杂合型),而反应体系中未加BSA的则无PCR扩增条带产生。结论 BSA可显著提高floxed Rb1基因的扩增效率,增加了该基因杂合子和纯合子的检出率并排除了假阴性结果,对于难以扩增的靶基因序列的PCR基因分型具有借鉴意义。Objective To investigate whether the addition of bovine serum albumin （BSA） into PCR system increase the efficiency and accuracy of floxed Rbl gene heterozygote and homozygote detection in osteosarcoma mouse models. Methods PCR genotyping of Rbl gene was performed using the offspring genome DNA of osteosarcoma mouse models as template. The used PCR primers contained the LoxP sites located in the two sides of exon 19 and the 3＇-end of Rbl gene. The genotyping results between PCR system added 0.16 mg/mL of BSA and not were compared. Results When BSA was added into the PCR system, the amplification products for the wild- type （showing a single 650 bp or 247 bp band）, homozygotic type （showing a single 700 bp or 295 bp band） and heterozygotic type （showing mixed bands of 650 bp and 700 bp or 247 bp and 295 bp） of RbI gene could be successfully obtained. However, when there was no BSA in the PCR system, no any amplification product was produced. Conclusion BSA may significantly increase the amplifi- cation efficiency of Rbl gene, and the detection rate of homozygotic and heterozygotic Rbl genes, which could avoid false negative re- suits, and would help to the PCR genotyping of target genes with DNA fragments difficult to be amplified.

关 键 词：小鼠骨肉瘤 RB1基因 PCR基因分型 牛血清清蛋白 

分 类 号：R733[医药卫生—肿瘤]