摘要
探讨mRNA长片段反转录PCR技术 (RT LDPCR)在cDNA芯片微量探针标记和信号放大中的应用。首先提取BEP2D细胞的总RNA ,然后用两种不同的方法进行标记 ,一种为RT LDPCR ,用荧光素Cy3 dCTP进行标记 ;另一种为传统的RNA反转录 ,用荧光素Cy5 dCTP进行标记。将两种方法标记好的探针等量混合后与含有 44 0个点 (4 4个基因 )的cDNA芯片同时杂交 ,发现二者具有很高的一致性 (0 .5 <Cy3 Cy5 >2 .0 )。由于RNA反转录法为cDNA芯片探针标记的传统方法 ,从而验证了RT LDPCR用于cDNA芯片探针标记的可行性。RT LDPCR具有对样品总RNA的需要量少和可对样品中信号进行放大的优点 ,特别适合于对材料来源受到限制的RNA进行标记。
This study aims at exploring the process of using reverse transcript coupling long distance PCR(RT LDPCR)to label and amplify trace probe. Firstly, total RNA from BEP2D cells was extracted and labeled by two different methods respectively, one was RT LDPCR using Cy3 dCTP as fluorescent dye; the other was traditional RNA reverse transcript using Cy5 dCTP as fluorescent dye. Then the probes labeled by these two methods were mixed equally and hybridized with the cDNA microarray. After scaning using ScanArray 3000, The high uniform of the two methods labeled probes was obtained and that proved the feasibility of using RT LDPCR to label cDNA microarray probe. The method of RT LDPCR can amplify the original probe signal, so it is very useful, especially when the RNA sample source is limited.
出处
《生物技术通讯》
CAS
2001年第4期254-256,共3页
Letters in Biotechnology
基金
国家重点基础研究发展规划 ( 973项目 )资助 (G19980 5 12 0 8)
