COMPARISON OF GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE AND 28S-RIBOSOMAL RNA GENE-EXPRESSION AS RNA LOADING CONTROLS FOR NORTHERN BLOT ANALYSIS OF CELL-LINES OF VARYING MALIGNANT POTENTIAL
COMPARISON OF GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE AND 28S-RIBOSOMAL RNA GENE-EXPRESSION AS RNA LOADING CONTROLS FOR NORTHERN BLOT ANALYSIS OF CELL-LINES OF VARYING MALIGNANT POTENTIAL
复制标题
DOI：
10.1006/abio.1994.1028
复制
发表时间：
1994-01-01
影响因子：
2.9
通讯作者：
WRIGHT, JA
中科院分区：
文献类型：
--
作者：
BHATIA, P;TAYLOR, WR;WRIGHT, JA
甘油醛-3-磷酸脱氢酶（GAPDH）已成为Northern blot分析中选择的基因，作为内部RNA装载控制。我们研究了GAPDH和28S-核糖体RNA （28S rRNA）基因在小鼠10T1 - 2细胞和从10T1 - 2细胞系衍生的多种致瘤性和高度恶性转移细胞系中的表达。我们观察到GAPDH mRNA水平在致瘤性和高度恶性细胞系之间存在显著差异，并且与正常小鼠10T1 2细胞相比，这些细胞系中的GAPDH mRNA水平升高。相比之下，28S rRNA的水平在致瘤性和高度恶性细胞系之间没有显著差异，与正常亲本小鼠10T1 2细胞系的水平大致相同。这些观察结果表明，当使用GAPDH基因表达作为Northern印迹的RNA负载控制时，应该非常谨慎。基于这些观察结果，我们建议使用28S rRNA基因表达作为Northern blot分析中使用总RNA的首选RNA负载控制。
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) has been a gene of choice in Northern blot analyses as an internal RNA loading control. We have investigated the expression of GAPDH and 28S-ribosomal RNA (28S rRNA) genes in mouse 10T1 2 cells and in a variety of tumorigenic and highly malignant metastatic cell lines derived from the 10T1 2 cell line. We observed that GAPDH mRNA levels varied markedly among the tumorigenic and highly malignant cell lines and were elevated in these cell lines when compared to the normal mouse 10T1 2 cells. In contrast, the levels of 28S rRNA did not significantly vary among the tumorigenic and highly malignant cell lines and were approximately at the same level as that found in the normal parental mouse 10T1 2 cell line. These observations indicate that much caution should be taken when using GAPDH gene expression as an RNA loading control for Northern blots. Based upon these observations, we recommend the use of 28S rRNA gene expression as a preferred RNA loading control for Northern blot analysis in which total RNA is used.