CN103498005A - Il-6基因pcr检测引物 - Google Patents

Il-6基因pcr检测引物 Download PDF

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CN103498005A
CN103498005A CN201310503149.7A CN201310503149A CN103498005A CN 103498005 A CN103498005 A CN 103498005A CN 201310503149 A CN201310503149 A CN 201310503149A CN 103498005 A CN103498005 A CN 103498005A
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primer
gene pcr
pcr detection
detection primer
detection
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唐余龙
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    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/158—Expression markers

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Abstract

本发明提供了IL-6基因PCR检测引物：正向引物(SEQIDNO:1)：5-GGCCCTTGCTTTCTCTTCG-3；反向引物（SEQIDNO:2：5-ATAATAAAGTTTTGATTATGT-3；用实时荧光PCR方法进行检测，得出IL-6的表达水平，作为判断免疫异常性疾病或急慢性炎症的判断依据，具有良好的特异性和实用性。

Description

IL-6基因PCR检测引物
 
技术领域
本发明属于生物技术检测领域，具体涉及IL-6因子基因检测技术。
背景技术
白细胞介素6（IL-6），主要由单核巨噬细胞、T细胞、血管内皮细胞、成纤维细胞产生，主要作用包括：刺激活化B细胞增殖，分泌抗体；活化T细胞增殖及CTL活化；刺激肝细胞合成急性期蛋白，参与炎症炎症反应；促进血细胞发育。IL-6是肝脏和急性期反应的重要调节因子。目前，研究发现的疾病中与IL-6异常表达有关，包括免疫异常性疾病：临床表现为发热、淋巴结肿大、血沉加快、急性期蛋白增高、高球蛋白白血症、自身抗体阳性等症状；亚急性甲状腺炎发作期间。另外，IL-6水平升高还与急慢性炎症、烧伤、移植排斥等相关。
目前，对IL-6血清水平的检测主要通过ELISA进行检测，但是本方法中使用特异性抗体价格较高，操作周期长，不适合对并病人治疗过程中IL-6指标实时定点检测。本发明提供一种IL-6基因检测引物，利用PCR检测技术定量和定性的检测IL-6基因表达，简单快捷，适合推广使用。
发明内容
为了解决上述现有技术的不足，本发明提供了IL-6基因PCR检测的引物。
本发明所采用的技术方案如下：
IL-6基因PCR检测引物：
正向引物(SEQ ID NO:1)：5-GGCCCTTGCTTTCTCTTCG-3
反向引物（SEQ ID NO:2：5-ATAATAAAGTTTTGATTATGT-3；
本发明的技术效果如下：
所用的引物针对IL-6的mRNA水平进行检测，提取血细胞的mRNA，并反转录为cDNA作为检测样品，运用实时荧光PCR方法进行检测，得出IL-6的表达水平，作为判断免疫异常性疾病或急慢性炎症的判断依据，具有良好的特异性和实用性。
附图说明
图1是实施例1的实时荧光PCR检测曲线。
具体实施方式
具体结合实施例对本发明作进一步的说明。
实施例1：
根据IL-6基因的cDNA进行设计PCR检测引物，引物序列如下：
正向引物(SEQ ID NO:1)：5-GGCCCTTGCTTTCTCTTCG-3
反向引物（SEQ ID NO:2：5-ATAATAAAGTTTTGATTATGT-3；
检测步骤：
1）提取mRNA：
   其中取免疫异常性疾病病人3例的血清，标记为1、2、3，取亚急性甲状腺炎的病人1例，标记4，取急性炎症病人2例，标记为5、6；阴性对照2例正常人血清。离心分离，并提取血液mRNA（全血mRNA提取试剂盒，罗氏公司），并反转录为cDNA， 
2）制备实时荧光PCR检测反应体系20微升：
取步骤1）中cDNA溶液2微升，加入10微升SYBR Premix Ex Taq(2×)、正向引物10μM、反向引物10μM、Rox  Dye（50×）0.4微升；并补加无菌水至20微升。
3）进行反应，反应温度如下：
预变性95℃ /100s，1 次循环；变性95℃ /30s，延伸66℃ /120s，40 次循环。
其中如附图1所示的PCR扩增曲线，其中以高于阴性对照的荧光0.245696设置阈值绘制PCR扩增曲线；图1的扩增曲线表明，病人1、2的实时荧光PCR曲线高于阴性对照和其他4个病例，3、4高于急性炎症病人。并且6位病例中的IL-6基因表达情况高于阴性对照。
                         SEQUENCE LISTING
 
<110>  余龙, 唐
 
<120>  IL-6基因PCR检测引物
 
<130>  2013
 
<160>  2    
 
<170>  PatentIn version 3.5
 
<210>  1
<211>  19
<212>  DNA
<213>  人工合成
 
<400>  1
ggcccttgct ttctcttcg                                                    19
 
 
<210>  2
<211>  21
<212>  DNA
<213>  人工合成
 
<400>  2
ataataaagt tttgattatg t                                                 21
 
 

Claims (1)

1.IL-6基因PCR检测引物，其特征是：
正向引物(SEQ ID NO:1)：5-GGCCCTTGCTTTCTCTTCG-3
反向引物（SEQ ID NO:2：5-ATAATAAAGTTTTGATTATGT-3。
CN201310503149.7A 2013-10-24 2013-10-24 Il-6基因pcr检测引物 Pending CN103498005A (zh)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105256038A (zh) * 2015-10-28 2016-01-20 中国人民解放军第二军医大学 裸鼹鼠细胞因子il-6基因pcr检测引物及检测试剂盒

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101608222A (zh) * 2008-06-20 2009-12-23 上海主健生物工程有限公司 糖尿病遗传检测试剂盒

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101608222A (zh) * 2008-06-20 2009-12-23 上海主健生物工程有限公司 糖尿病遗传检测试剂盒

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
高文 等: "辛伐他汀对血管紧张素II刺激人单核细胞分泌白介素6的影响", 《临床荟萃》, vol. 24, no. 11, 5 June 2009 (2009-06-05), pages 913 - 934 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105256038A (zh) * 2015-10-28 2016-01-20 中国人民解放军第二军医大学 裸鼹鼠细胞因子il-6基因pcr检测引物及检测试剂盒

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