RT-PCR-based analysis of microRNA (miR-1 and-124) expression in mouse CNS

被引:128
作者
Mishima, Takuya
Mizuguchi, Yoshiaki
Kawahigashi, Yutaka
Takizawa, Takami
Takizawa, Toshihiro
机构
[1] Nippon Med Sch, Dept Mol Anat & Cell Biol, Tokyo 1138602, Japan
[2] Nippon Med Sch, Dept Surg Organ Funct & Biol Regulat, Tokyo 1138602, Japan
关键词
microRNA; miR-1; miR-124; real-time PCR; differential expression; central nervous system;
D O I
10.1016/j.brainres.2006.11.035
中图分类号
Q189 [神经科学];
学科分类号
071006 ;
摘要
More than 700 microRNAs (miRNAs) have been cloned, and the functions of these molecules in developmental timing, cell proliferation, and cancer have been investigated widely. MiRNAs are analyzed with Northern blot and sequential colony evaluation; however, reverse transcription-polymerase chain reaction (RT-PCR)-based miRNA assay remains to be developed. In this report, we describe improved real-time RT-PCR methods using specific or non-specific RT primer for the semi-quantitative analysis of miRNA expression. The use of the new methods in a model study revealed differential expression of miRNA-1 (miR-1) and miR-124 in mouse organs. Specifically, our methods revealed that miR-124 concentrations in the mouse central nervous system (CNS; cerebral cortex, cerebellum, and spinal cord) were more than 100 times those in other organs. By contrast, miR-1 expression in the CNS was 100-1000 times lower than that in skeletal muscle and heart. Furthermore, we revealed anatomically regional differences in miR-124 expression within the CNS: expression ratios versus the cerebral cortex were 60.7% for the cerebellum and 35.4% for the spinal cord. These results suggest that our RT-PCR-based methods would be a powerful tool for studies of miRNA expression that is associated with various neural events. (c) 2006 Elsevier B.V. All rights reserved.
引用
收藏
页码:37 / 43
页数:7
相关论文
共 16 条
[1]   Quantitation of microRNAs using a modified Invader assay [J].
Allawi, HT ;
Dahlberg, JE ;
Olson, S ;
Lund, E ;
Olson, M ;
Ma, WP ;
Takova, T ;
Neri, BP ;
Lyamichev, VI .
RNA, 2004, 10 (07) :1153-1161
[2]   bantam encodes a developmentally regulated microRNA that controls cell proliferation and regulates the proapoptotic gene hid in Drosophila [J].
Brennecke, J ;
Hipfner, DR ;
Stark, A ;
Russell, RB ;
Cohen, SM .
CELL, 2003, 113 (01) :25-36
[3]   A microRNA polycistron as a potential human oncogene [J].
He, L ;
Thomson, JM ;
Hemann, MT ;
Hernando-Monge, E ;
Mu, D ;
Goodson, S ;
Powers, S ;
Cordon-Cardo, C ;
Lowe, SW ;
Hannon, GJ ;
Hammond, SM .
NATURE, 2005, 435 (7043) :828-833
[4]  
Hipfner DR, 2002, GENETICS, V161, P1527
[5]   Identification of novel genes coding for small expressed RNAs [J].
Lagos-Quintana, M ;
Rauhut, R ;
Lendeckel, W ;
Tuschl, T .
SCIENCE, 2001, 294 (5543) :853-858
[6]   An abundant class of tiny RNAs with probable regulatory roles in Caenorhabditis elegans [J].
Lau, NC ;
Lim, LP ;
Weinstein, EG ;
Bartel, DP .
SCIENCE, 2001, 294 (5543) :858-862
[7]   An extensive class of small RNAs in Caenorhabditis elegans [J].
Lee, RC ;
Ambros, V .
SCIENCE, 2001, 294 (5543) :862-864
[8]   THE C-ELEGANS HETEROCHRONIC GENE LIN-4 ENCODES SMALL RNAS WITH ANTISENSE COMPLEMENTARITY TO LIN-14 [J].
LEE, RC ;
FEINBAUM, RL ;
AMBROS, V .
CELL, 1993, 75 (05) :843-854
[9]   Microarray analysis shows that some microRNAs downregulate large numbers of target mRNAs [J].
Lim, LP ;
Lau, NC ;
Garrett-Engele, P ;
Grimson, A ;
Schelter, JM ;
Castle, J ;
Bartel, DP ;
Linsley, PS ;
Johnson, JM .
NATURE, 2005, 433 (7027) :769-773
[10]  
Lu David P, 2005, J RNAi Gene Silencing, V1, P44