Real-time quantitative PCR（qRT-PCR） technology has the advantages of high sensitivity， strong specificity， good repeatability， and accurate quantification. It is widely used in quantitative analysis of target gene expression， and screening of internal reference genes with stable expression levels is improved an important prerequisite for the accuracy of qRT-PCR results， Lactobacillus bulgaricus is not only a common strain of yogurt starter， but also the main strain that causes acidification after yogurt. In order to select the qRT-PCR internal reference gene for expression analysis of functional genes related to acidification after Lactobacillus bulgaricus， the model strain ATCC 11842 of Lactobacillus bulgaricus was used as the test strain. Five candidate internal reference genes （16SrRNA， rpoB， ldh， rodA， recA）， using qRT-PCR technology， studied the expression level of the candidate internal reference gene under acid stress and acid cold stress， that is， Ct value changes； using three software geNorm， NormFinder and Bestkeeper， analyzed and compared the expression stability of qRT-PCR of candidate internal reference genes under acid stress and acid cold stress， and screened the most suitable internal reference genes for qRT-PCR； using the optimal internal reference genes obtained by screening， analysis the relative expression levels of qRT-PCR of three target genes（poxI， Ldb1301， dnaJ） of ATCC 11842 strain under acid stress and acid cold stress were verified， and the reliability of the internal reference genes obtained by screening was verified. The results show that the Ct value of the same candidate internal reference gene under acid stress and acid cold stress has the smallest change in the expression of rpoB and recA； the results of the analysis and comparison of the three softwares are consistent： rpoB and recA are the most expressed under acid stress and acid cold stress two stable genes， the most suitable qRT-PCR selected internal reference genes are rpoB and recA； the analysis of the relative expression of qRT-PCR under three acid genes（poxI， Ldb1301， dnaJ） under acid stress and acid cold stress the results were consistent with the results of the previous transcriptomics sequencing analysis， which further confirmed the reliability of the two internal reference genes rpoB and recA obtained in this study. This article provides a reliable basis for in-depth research on the expression of post-acidification function genes of Lactobacillus bulgaricus using qRT-PCR technology， and further reveals the post-acidification mechanism and selective breeding of weak post-acidification strains； functional gene expression under acidification or under different conditions provides guidance.