无花果RNA的提取和半定量RT-PCR内参基因的优选

RNA isolation and internal reference gene selection for semi-quantitative RT-PCR of fig(Ficus carica)

本试验以无花果的根、茎、叶、果作为材料,比较了CTAB法和Trizol法对无花果材料RNA的提取效果。以CTAB法提取的不同无花果材料的RNA为模板,反转成cDNA第一链,通过半定量RT-PCR,研究了植物常用内参基因18SrRNA、Actin和Tubulin的表达量变化。结果表明:CTAB法是适合不同无花果材料的RNA提取法;18SrRNA在无花果不同组织中的表达水平较高,且相对稳定,Tubulin在无花果不同组织中相对表达量较低,且相对稳定,是研究无花果不同组织基因表达水平较为适宜的内参基因。

Fig is an ancient fruit tree species with important economic value.Along with the development of fruit tree molecule biology,there are more and more such studies on figs,and one important area is the expression level of functional genes.RNA isolation and internal reference gene selection for semi-quantitative RT-PCR are key steps in gene expression studies.In this paper,fig root,stem,young leaf and young fruit were sampled.RNA isolation methods,i.e.Trizol and CTAB,were compared for their RNA extraction results.RNA quality results indicated that CTAB method had better performance in fig RNA isolation.Using fig RNA extracted with CTAB method from different tissues,after reversing transcription and the gain of the first strand cDNA,we studied the expression of three house-keeping genes to evaluate their feasibility as internal reference for semi-quantitative RT-PCR.Among of these three genes,18S rRNA showed rather high and stable expression level in four different tissues,while Tubulin revealed relatively low but stable expressions.The expression of Actin demonstrated differentiated expression in four different organs.Thus,18S rRNA and Tubulin were suggested as internal references.This study lays the ground for future study of gene expression in fig by semi-quantitative RT-PCR.