Leder Erica H, Merilä Juha, Primmer Craig R
Division of Genetics and Physiology, Department of Biology (Vesilinnantie 5), University of Turku, Turku FI-20014, Finland.
BMC Genomics. 2009 Sep 11;10:426. doi: 10.1186/1471-2164-10-426.
The use of microarray technology for describing changes in mRNA expression to address ecological and evolutionary questions is becoming increasingly popular. Since three-spine stickleback are an important ecological and evolutionary model-species as well as an emerging model for eco-toxicology, the ability to have a functional and flexible microarray platform for transcriptome studies will greatly enhance the research potential in these areas.
We designed 43,392 unique oligonucleotide probes representing 19,274 genes (93% of the estimated total gene number), and tested the hybridization performance of both DNA and RNA from different populations to determine the efficacy of probe design for transcriptome analysis using the Agilent array platform. The majority of probes were functional as evidenced by the DNA hybridization success, and 30,946 probes (14,615 genes) had a signal that was significantly above background for RNA isolated from liver tissue. Genes identified as being expressed in liver tissue were grouped into functional categories for each of the three Gene Ontology groups: biological process, molecular function, and cellular component. As expected, the highest proportions of functional categories belonged to those associated with metabolic functions: metabolic process, binding, catabolism, and organelles.
The probe and microarray design presented here provides an important step facilitating transcriptomics research for this important research organism by providing a set of over 43,000 probes whose hybridization success and specificity to liver expression has been demonstrated. Probes can easily be added or removed from the current design to tailor the array to specific experiments and additional flexibility lies in the ability to perform either one-color or two-color hybridizations.
利用微阵列技术描述mRNA表达变化以解决生态和进化问题正变得越来越流行。由于三刺鱼是一种重要的生态和进化模式物种，也是生态毒理学的新兴模式生物，拥有一个功能强大且灵活的微阵列平台用于转录组研究将极大地提升这些领域的研究潜力。
我们设计了代表19274个基因（占估计基因总数的93%）的43392个独特寡核苷酸探针，并测试了来自不同种群的DNA和RNA的杂交性能，以确定使用安捷伦阵列平台进行转录组分析的探针设计效果。大多数探针是有功能的，这从DNA杂交成功得到证明，并且30946个探针（14615个基因）对从肝脏组织分离的RNA有显著高于背景的信号。在肝脏组织中被鉴定为表达的基因被归入三个基因本体论组（生物过程、分子功能和细胞组分）中的每一个的功能类别。正如预期的那样，功能类别中比例最高的属于那些与代谢功能相关的类别：代谢过程、结合、分解代谢和细胞器。
本文提出的探针和微阵列设计通过提供一组超过43000个探针，其杂交成功以及对肝脏表达的特异性已得到证明，为这一重要研究生物体的转录组学研究提供了重要一步。探针可以很容易地从当前设计中添加或移除，以根据特定实验定制阵列，并且额外的灵活性在于能够进行单色或双色杂交。
