Postier Bradley, Didonato Raymond, Nevin Kelly P, Liu Anna, Frank Bryan, Lovley Derek, Methe Barbara A
Department of Microbiology, University of Massachusetts Amherst, MA 01003, USA.
J Microbiol Methods. 2008 Jul;74(1):26-32. doi: 10.1016/j.mimet.2007.07.004. Epub 2007 Jul 17.
Although the genome sequences of many microorganisms are now known, whole-genome DNA microarray platforms consisting of PCR amplicon, or oligonucleotide elements printed onto glass slides have been readily available for only a relatively few, highly studied microorganisms. For those microorganisms more recently cultured or studied by fewer investigators it has been difficult to justify the initial time and expense of developing such array platforms especially if only a limited number of gene expression studies are envisioned. However, in-situ synthesized oligonucleotide (ISO) arrays can be inexpensively fabricated on an 'as needed' basis with a reduced initial investment in time, personnel, resources, and costs. To evaluate the performance of one ISO array platform, gene expression patterns in Geobacter sulfurreducens under nitrogen-fixing conditions were compared with results from quantitative reverse transcriptase PCR (qRT-PCR) and previously published data from a similar experiment using spotted PCR amplicon arrays. There were strong correlations between the results of the ISO arrays and the results from qRT-PCR (r(2)=0.762) and spotted array (r(2)=0.744) analyses. After initial use the ISO arrays could be successfully stripped and reused. The increased flexibility in array design and reusability coupled with a lower initial investment in terms of fabrication time and cost for the ISO arrays suggest that they may be the preferred approach when investigating gene expression in microorganisms, especially when only a few expression studies are required.
尽管现在已经知晓许多微生物的基因组序列,但由印在载玻片上的PCR扩增子或寡核苷酸元件组成的全基因组DNA微阵列平台,仅适用于相对较少的、经过深入研究的微生物。对于那些最近才被培养或研究人员较少研究的微生物来说,开发这样的阵列平台所需的初始时间和费用很难说得过去,尤其是如果预计只有有限数量的基因表达研究的话。然而,原位合成寡核苷酸(ISO)阵列可以根据“按需”原则廉价制造,在时间、人员、资源和成本方面的初始投资较少。为了评估一种ISO阵列平台的性能,将固氮条件下嗜硫地杆菌的基因表达模式与定量逆转录PCR(qRT-PCR)的结果以及先前发表的使用斑点PCR扩增子阵列的类似实验数据进行了比较。ISO阵列的结果与qRT-PCR(r² = 0.762)和斑点阵列(r² = 0.744)分析的结果之间存在很强的相关性。初次使用后,ISO阵列可以成功剥离并重复使用。ISO阵列在阵列设计和可重复使用性方面具有更大的灵活性,加上在制造时间和成本方面的初始投资较低,这表明在研究微生物基因表达时,尤其是在只需要进行少数表达研究的情况下,它们可能是首选方法。