Grup de Recerca en Biotecnologia Enològica, Departament de Bioquímica i Biotecnologia, Facultat d'Enologia de Tarragona, Universitat Rovira i Virgili, Tarragona, Spain.
J Appl Microbiol. 2009 Feb;106(2):666-74. doi: 10.1111/j.1365-2672.2008.04046.x.
The aim of this study was to develop a reliable system to analyse the expression of the pyrroloquinoline quinone (PQQ)-alcohol dehydrogenase (ADH) and test its ability to predict the growth and oxidative activity of some acetic acid bacteria (AAB).
Specific primers were designed for use in RT-PCR to quantify ADH expression and several housekeeping genes in four species of AAB. 16S rRNA gene was selected as an internal control. The relative expression of adhA was measured in Acetobacter aceti, Acetobacter pasteurianus, Gluconacetobacter hansenii and Gluconobacter oxydans grown in two media that had glucose or ethanol as the carbon source. AAB adhA expression was shown to be related to the two Acetobacter species' ability to oxidise and grow on ethanol, whereas G. oxydans were unable to grow on ethanol and the growth of Ga. hansenii was not related to adhA expression.
The differential expression of ADH could be a marker to analyse both growth and oxidation ability in some AAB, especially those of the genus Acetobacter.
Several housekeeping genes were tested in AAB and after growth in different media and it was evident that only the ribosomal coding genes were adequate as reference genes for RT-PCR.
本研究旨在开发一种可靠的系统来分析吡咯并喹啉醌(PQQ)-醇脱氢酶(ADH)的表达,并测试其预测某些醋酸菌(AAB)生长和氧化活性的能力。
设计了用于 RT-PCR 的特异性引物,以定量四种 AAB 中 ADH 表达和几种管家基因。16S rRNA 基因被选为内参。在以葡萄糖或乙醇为碳源的两种培养基中,测量了在醋酸杆菌、巴氏醋酸杆菌、葡萄糖醋酸杆菌和氧化葡萄糖酸杆菌中生长的 adhA 的相对表达。结果表明,AAB adhA 的表达与两种醋酸杆菌氧化和利用乙醇的能力有关,而 G. oxydans 不能利用乙醇生长,Ga. hansenii 的生长与 adhA 表达无关。
ADH 的差异表达可以作为分析某些 AAB 生长和氧化能力的标志物,特别是醋酸杆菌属。
在不同的 AAB 中测试了几种管家基因,并在不同的培养基中生长后,显然只有核糖体编码基因适合作为 RT-PCR 的参考基因。