CN109628570A - 一种检测TRIM29基因Tyr544Cys突变的试剂盒及其用途 - Google Patents

一种检测TRIM29基因Tyr544Cys突变的试剂盒及其用途 Download PDF

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CN109628570A
CN109628570A CN201811489797.0A CN201811489797A CN109628570A CN 109628570 A CN109628570 A CN 109628570A CN 201811489797 A CN201811489797 A CN 201811489797A CN 109628570 A CN109628570 A CN 109628570A
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trim29
mutation
kit
tyr544cys
detection
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侯金林
蒋德科
周彬
孙剑
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Southern Hospital Southern Medical University
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Abstract

本发明属于医疗领域，公开了一种检测TRIM29基因Tyr544Cys突变的试剂盒，该试剂盒可预测慢性乙型肝炎核苷(酸)类似物抗病毒治疗应答的疗效。该试剂盒含有针对TRIM29特异性扩增的引物。所述的引物包括RM2.F和RM2.R；所述的RM2.F的序列为CATGGCCCAGAGAAGTCCT；所述的RM2.R序列为GGTGGTCACTTTGGAGAAGG。本发明的目的在于提供一种检测TRIM29基因Tyr544Cys突变的试剂盒，其优点在于有利于进一步认识慢性乙型肝炎应用核苷(酸)类似物抗病毒治疗应答的疗效预测的发生机理，并通过在临床上开展治疗前的TRIM29突变筛查工作，对慢性乙型肝炎核苷(酸)类似物抗病毒治疗应答的疗效进行预测。

Description

一种检测TRIM29基因Tyr544Cys突变的试剂盒及其用途
技术领域
本发明涉及医疗领域，具体涉及一种检测TRIM29基因Tyr544Cys突变的试剂盒及其对慢性乙型肝炎抗病毒治疗应答疗效的预测。
背景技术
CN 201110041465.8公开了一种预测干扰素治疗慢性乙型肝炎疗效的血浆miRNA谱及检测试剂盒，该试剂盒含有针对TRIM29特异性变异的引物。
预测慢性乙型肝炎疗效的检测方法及检测试剂盒，该试剂盒包含一对特异性扩增的引物。同时提供了相应的、高效、低廉的试剂盒和检测方法。其检测患者基因组的TRIM29应答相关SNP位点，适合应用于临床判断核苷(酸)类似物治疗的有效性。与其他已知与疗效相关因素相比，此位点是一个独立的预测因素。将此SNP位点与病人治疗前其他指标结合可获得更高的预测准确率。此位点与HBV相关其他临床指标结合将有助于个体化治疗方案的开展，最终降低治疗费用并提高完全应答的比例。
在实际应用过程中，如果有更多的辅助手段来进行相应的疗效确定，则可靠性更高。
发明内容
本发明的目的是提供一种慢性乙型肝炎抗病毒治疗应答疗效预测的引物、试剂盒，其优点在于有利于进一步认识影响慢性乙型肝炎抗病毒治疗应答的因素，并通过在临床上开展治疗前的TRIM29突变筛查工作，对慢性乙型肝炎抗病毒治疗应答的疗效进行预测。
本发明的具体方案如下：一种慢性乙型肝炎抗病毒治疗应答疗效预测的引物，所述的引物包括RM2.F和RM2.R；所述的RM2.F的序列为CATGGCCCAGAGAAGTCCT；所述的RM2.R序列为GGTGGTCACTTTGGAGAAGG。
同时本发明还公开了一种慢性乙型肝炎抗病毒治疗应答疗效预测的试剂盒，所述的试剂盒的引物如上所述，其为针对TRIM29特异性扩增的引物。
本发明的有益效果为：
TRIM29突变基因，其含有TRIM29基因序列，并且还含有位于TRIM29基因1631位核苷酸A→G杂合或纯合突变；本发明从待测样品中得到核苷酸序列与TRIM29正常基因的序列进行比较，核对突变是否为1631位核苷酸A→G的杂合或纯合突变，以及按照正常读码框架进行翻译以确定是否存在Tyr544Cys氨基酸突变位点；本发明还公开了通过测序检测所述TRIM29突变基因的试剂盒。本发明所述TRIM29突变基因及检测方法将有利于进一步认识影响慢性乙型肝炎抗病毒治疗应答疗效的因素，并通过在临床上开展治疗前的TRIM29突变筛查工作，对慢性乙型肝炎抗病毒治疗应答的疗效进行预测。
附图说明
图1为实施例1的电泳结果图。
图2-1、图2-2、图2-3为实施例1的扩增得到的三份样品的测序结果图。
具体实施方式
下面结合具体实施方式，对本发明的技术方案作进一步的详细说明，但不构成对本发明的任何限制。
实施例1：
提取待测全血中的DNA
应用引物扩增DNA片段
引物序列：
RM2.F:CATGGCCCAGAGAAGTCCT
RM2.R:GGTGGTCACTTTGGAGAAGG
体系配置:
PCR循环参数如下
其中10X LABuffer，dNTPs和LA-taq从TARARA公司购买，引物在华大基因公司合成
PCR产物检测：
用1.5％琼脂糖凝胶对PCR扩增产物进行电泳，以检测其是否为目的片段并确认其纯度。PCR产物经电泳显示单一条带，无杂带，则提示PCR产物成分单一，无非特异性扩增，电泳带位置位于适当大小的位置，则为目的片段。根据上述引物扩增目的片段应为287bp.
PCR产物的纯化和测序：
PCR产物经切胶回收目的片段后用ABI3730测序仪进行测序。
将得到的序列与TRIM29正常基因组的标准序列(NCBI Accession number)利用MEGA7软件进行比对确定TRIM29是否含有1631位核苷酸A→G杂合或纯合突变。电泳结果图如图1所示，其中标为5,6,25的泳道是编号为5,6,25的3份样本经过引物扩增后获得的目的条带，标为“-”的泳道是阴性对照，标为M的泳道为DNA markerⅢ(购自天根生化科技有限公司)，电泳结果显示3份样本目的条带位于200-500bp之间，符合目的条带为287bp的预判。目的条带清晰，没有非特异性扩增，本扩增方法效率高，结果可靠；测序结果如图2所示，图2-1，2，3分别为以上扩增得出的三份样本送测序公司测序后获得的测序峰图，可以看出不同的样本在1631位核苷酸为不同的SNP，其中图2-1的样本在该位置为G纯合子，图2-2的样本在该位置为GA杂合子。图2-3的样本在该位置为A纯合子。
上述实施例为本发明较佳的实施方式，但本发明的实施方式并不受上述实施例的限制，其它的任何未背离本发明的精神实质与原理下所作的改变、修饰、替代、组合、简化，均应为等效的置换方式，都包含在本发明的保护范围之内。
SEQUENCE LISTING
<110> 南方医科大学南方医院
<120> 一种检测TRIM29基因Tyr544Cys突变的试剂盒及其用途
<130>
<160> 1
<170> PatentIn version 3.3
<210> 1
<211> 19
<212> DNA
<213> 未知
<400> 1
catggcccag agaagtcct 19
SEQUENCE LISTING
<110> 南方医科大学南方医院
<120> 一种检测TRIM29基因Tyr544Cys突变的试剂盒及其用途
<130>
<160> 1
<170> PatentIn version 3.3
<210> 1
<211> 20
<212> DNA
<213> 未知
<400> 1
ggtggtcact ttggagaagg 20

Claims (3)

1.检测TRIM29基因Tyr544Cys突变的试剂盒，其特征在于，含有针对TRIM29特异性扩增的引物，所述的引物包括RM2.F和RM2.R；所述的RM2.F的序列为CATGGCCCAGAGAAGTCCT；所述的RM2.R序列为GGTGGTCACTTTGGAGAAGG。
2.一种检测TRIM29基因Tyr544Cys突变的试剂盒，其用于预测核苷(酸)类似物治疗慢性乙型肝炎疗效。
3.一种预测核苷(酸)类似物治疗慢性乙型肝炎疗效的试剂盒，其特征在于，含有如权利要求1所述针对TRIM29特异性扩增的引物。
CN201811489797.0A 2018-12-07 2018-12-07 一种检测TRIM29基因Tyr544Cys突变的试剂盒及其用途 Pending CN109628570A (zh)

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