L-乳酸脱氢酶基因在乳酸乳球菌KLDS4.0325中的表达

Variations in the Expression of the L-lactate Dehydrogenase Gene during the Different Phases of Lactococcus lactis KLDS4.0325 Growth

本文研究了在m RNA转录水平上三种L-乳酸脱氢酶基因在乳酸乳球菌KLDS4.0325不同生长阶段的表达情况。以16s r RNA作为内参基因,应用实时荧光定量PCR技术测定不同生长阶段的菌体中L-乳酸脱氢酶基因(ldh、ldh X和ldh B)表达的动态变化规律。该菌株的生长曲线呈S型,培养2~6 h为菌体的生长对数期,随后进入稳定期。生长曲线的测定表明该菌株生长力极其旺盛;16s r RNA、ldh、ldh X和ldh B基因的扩增效率曲线显示,内参基因与目的基因的扩增效率一致且接近100%,这说明利用相对荧光定量法能够较好的反应目的基因的表达量;ldh、ldh X和ldh B基因在菌体生长的3个时间点(6 h、9 h和12 h)都有一定量的表达,随着菌体的不断生长,基因表达的变化均呈现显著性上升趋势,且上升幅度不断增加,在12 h时基因表达量达到最大值。其中ldh基因在的表达量在菌体生长的3个时间点存在显著性差异,而ldh X、ldh B基因的表达量存在极显著差异。

The expressions of three L-lactate dehydrogenase genes at the m RNA transcription level during the different phases of Lactococcus lactis KLDS4.0325 growth were investigated in this study. Dynamic variations in the expressions of the three L-lactate dehydrogenase genes(ldh, ldh B, and ldh X) during the different growth phases were detected by fluorescence-based quantitative real time polymerase chain reaction(PCR), using the 16 s r RNA gene as the internal standard. The growth curves of the strain were observed to be ‘S'-shaped, with the strains reaching the exponential phase of growth within 2~6 h of culture; subsequently, the cells remained in the stationary phase. The growth curve revealed the extremely strong vitality of the cells; the amplification efficiency curve of the 16 s r RNA, ldh, ldh B, and ldh X genes indicated similar amplification efficiencies(close to 100%) of the reference and target genes. These results indicated that the fluorescence-based quantitative real time PCR method reflected the expression level of the target genes adequately. The ldh, ldh B, and ldh X genes were expressed to specific levels during the three time points(6 h, 9 h, and 12 h) of strain growth. In addition to the growth of the strain, the gene expression levels demonstrated a significant upward trend, with a continuous amplitude of increase; the highest expression level was observed at 12 h post-initiation. The levels of expression of the ldh gene significantly differed among the three time points; in addition, the expression of ldh B significantly differed from that of ldh X.