CN111349705A - circASXL1作为肺癌诊断标志物及其运用 - Google Patents
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Abstract
本发明公开了一种circASXL1作为肺癌诊断标志物及其应用，包含有ASXL1基因的环状RNA，设计并合成出特异性用于实时定量PCR的检测引物。针对目标人群对肺癌筛查依困难，以及临床上确诊的肺癌绝大多数都是中晚期的现状提出，在肺癌进程中找到分子预警信号，本发明发现circASXL1分子标志物肺癌标志物，包含1个试剂盒，可实现肺癌患者与非肺癌患者样本的区分，解决临床上早期肺癌癌难以及时、准确诊断的缺陷。
Description
技术领域
本发明属于生物标志物领域，具体地本发明涉及一种circASXL1作为肺癌诊断标志物及其应用。该标志可以很好的进行肿瘤的诊断。
背景技术
那些不编码蛋白质的RNA称为ncRNA调控ncRNA分为lncRNA和sncRNA。LncRNA可以是线性的也可以是环状的。环状RNA最早是在1976.3年发现的从那时起，已经发现了多种circRNA。CircRNA比其他lncRNA对核酸外切酶的抵抗力更强，更稳定。 同时，最近的研究表明， circRNA可以分为非编码circRNA和编码circRNA。 已发现许多circRNA发挥肿瘤抑制作用会在几种类型的癌症中起作用。 其中一些由单个外显子产生但是，大多数circRNA由多个外显子产生。circRNA与细胞的生长增殖分化等功能密切相关，在肿瘤的发生治疗中起重要作用。
发明内容
本发明的目的在于提供一种肺癌标志物在制备肺癌诊断产品中的应用，其标志物为ASXL1基因的环状RNA。其环状RNA的序列为SEQ ID NO.1所示，检测试剂为实时荧光定量PCR试剂盒，剂盒包含有序列如TCGCATGCCTCAATGCTATG （SEQ ID NO.2）和GTCCCACTTCTCATTTCCTTTAG（SEQ ID NO.3）所示的引物对。判断是否为肺癌采用ΔCt值。
本研究受国家自然科学基金的支持（81860572）。
本发明（优点）：方法非常简单可以通过提取RNA后进行荧光定量检测从而判定基因表达，整个过程仅仅需要2-3个小时，组织或者外周血需要量都很少，成本低，器材简单。
具体实施方式
下面对本发明作进一步的说明，但不以任何方式对本发明加以限制，基于本发明教导所作的任何变换或替换，均属于本发明的保护范围。
本发明所述的肺癌标志物为ASXL1基因的环状RNA。
所述的环状RNA的序列为SEQ ID NO.1所示。
检测序列为SEQ ID NO.1的环状RNA的检测试剂为实时荧光定量PCR试剂盒。
实时荧光定量PCR试剂盒包含有序列如SEQ ID NO.2和SEQ ID NO.3所示的引物对。
本发明所述的肺癌标志物的应用为所述的肺癌标志物在制备肺癌诊断产品中的应用。
实施例1
在取得知情同意后，收集15名确诊的肺癌患者的肺癌组织以及15名非肺癌但是需要手术的患者取正常组织10mg-100mg，使用TRIZOL提取RNA，采用逆转录试剂进行逆转录实时荧光定量PCR反应体系(15. 0μl ) 包括, 7.5μl 荧光定量试剂，1μl基因组DNA,1.5μl 引物上下游引物为（5μlmo l/L）, 去离子水5μl。PCR 反应条件为95 ℃ 10min预变性，95 ℃10s、60 ℃ 10 s、72℃ 10 s 40个循环得到Ct值，每个反应5个副孔，增加数据确定性。使用β-actin作用内参，circASXL1的表达公式。20-（Ct片段-Ct内参。）circASXL1上游引物：TCGCATGCCTCAATGCTATG (SEQ ID NO.2)下游引物： GTCCCACTTCTCATTTCCTTTAG (SEQ IDNO.3)，β-actin上游引物： GCCGAGGACTTTGATTGCAC (SEQ ID NO.4)下游引物：TGGACTTGGGAGAGGACTGG (SEQ ID NO.5)。结果肺癌组织表达为10.06，8.93，8.89，8.24，8.24，7.93，7.88，7.29，6.81，6.79，6.73，5.64，5.22，3.34，1.58，正常组织表达为14.02，14.01，13.83，13.05，12.27，11.98，11.92，11.5，11.46，11.36，11.33，11.22，10.16，9.76，9.22。将诊断值定为10.10可以较好分辨其是否为肺癌或者非肺癌患者。
SEQUENCE LISTING
<110> 昆明医科大学
<120> circASXL1作为肺癌诊断标志物及其运用
<130> 1
<140> 1
<141> 2020-01-15
<160> 5
<170> PatentIn version 3.3
<210> 1
<211> 195
<212> RNA
<213> 人类（Homo sapiens）
<400> 1
guauuagaaa acuacucgga ugcuccaaug acaccaaaac agauucugca ggucauagag 60
gcagaaggac uaaaggaaau gagaaguggg acuuccccuc ucgcaugccu caaugcuaug 120
cuacauucca auucaagagg aggagagggg uuguuuuaua aacugccugg ccgaaucagc 180
cuuuucacgc ucaag 195
<210> 2
<211> 20
<212> DNA
<213> 人工序列（Artificial sequence）
<400> 2
tcgcatgcct caatgctatg 20
<210> 3
<211> 23
<212> DNA
<213> 人工序列(Artificial sequence)
<400> 3
gtcccacttc tcatttcctt tag 23
<210> 4
<211> 20
<212> DNA
<213> 人工序列(Artificial sequence)
<400> 4
gccgaggact ttgattgcac 20
<210> 5
<211> 20
<212> DNA
<213> 人工序列(Artificial sequence)
<400> 5
tggacttggg agaggactgg 20
Claims (5)
1.一种肺癌标志物，其特征在于所述的标志物为ASXL1基因的环状RNA。
2.根据权利要求1所述的肺癌标志物，其特征在于所述的环状RNA的序列为SEQ IDNO.1所示。
3.根据权利要求2所述的肺癌标志物，其特征在于检测序列为SEQ ID NO.1的环状RNA的检测试剂为实时荧光定量PCR试剂盒。
4.根据权利要求3所述的肺癌标志物，其特征在于实时荧光定量PCR试剂盒包含有序列如SEQ ID NO.2和SEQ ID NO.3所示的引物对。
5.一种权利要求1~4任一所述的肺癌标志物的应用，其特征在于所述的肺癌标志物在制备肺癌诊断产品中的应用。
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111118011A (zh) * | 2020-02-11 | 2020-05-08 | 昆明医科大学 | 一种抑制hsa_circ_0027478表达的siRNA及其应用 |
| CN111118012A (zh) * | 2020-02-11 | 2020-05-08 | 昆明医科大学 | 一种抑制hsa_circ_0051680表达的siRNA及其应用 |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111118011A (zh) * | 2020-02-11 | 2020-05-08 | 昆明医科大学 | 一种抑制hsa_circ_0027478表达的siRNA及其应用 |
| CN111118012A (zh) * | 2020-02-11 | 2020-05-08 | 昆明医科大学 | 一种抑制hsa_circ_0051680表达的siRNA及其应用 |
| CN111118011B (zh) * | 2020-02-11 | 2022-09-06 | 昆明医科大学 | 一种抑制hsa_circ_0027478表达的siRNA及其应用 |
| CN111118012B (zh) * | 2020-02-11 | 2022-09-06 | 昆明医科大学 | 一种抑制hsa_circ_0051680表达的siRNA及其应用 |
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