Identification of genes differentially expressed in 1 dpa and 10 dpa fiber

A method was developed to isolate RNA from 1 dpa fiber. ESTs derived from this and other cotton mRNAs were sequenced and assembled into contigs. Contigs composed of EST unique to libraries of interest and unique singletons were represented on a microarray. Microarrays were hybridized with labed nucleic acids derived from 10 dpa fiber and 1 dpa fiber to identify genes differentially regulated during fiber initiation and fiber elongation. Genes with known expression in fiber were included to validate the microarray. Analysis of the gene ontology (GO) of differentially expressed genes suported previously reported aspects of fiber initiation such as cell wall remodeling. Transcription factors upregulated in fiber initials and elongating fiber were identified Keywords: cell type comparison The microarray represents 11143 gene models. One gene model is represented by 5 different oligonucleotides, 8604 gene models are represented by 4 different oligos, 1706 gene models are represented by 3 different oligonucleotides, 831 gene models are represented by 3 of the same oligonucleotides and one gene model is represented by 2 different oligonucleotides. Each micro array is hybridized with a labeled nucleic acid derived form RNA isolated from 1 dpa fiber or 10 dpa fiber harvested on different days from field grown plants. One is a dye swap.

本研究开发了一种从开花后1天（days post anthesis, 简称dpa）棉纤维中提取RNA的方法。由此棉纤维RNA及其他棉花mRNA获得的表达序列标签（Expressed Sequence Tags, EST）经测序并组装为重叠群（contigs）。由目标文库特异性EST与独特单序列（singletons）构成的重叠群被制备至基因芯片（microarray）上。将该基因芯片与取自10 dpa和1 dpa棉纤维的标记核酸进行杂交，以鉴定在棉纤维起始与伸长过程中受差异调控的基因。纳入已知在棉纤维中表达的基因，以验证基因芯片实验的可靠性。对差异表达基因的基因本体论（Gene Ontology, GO）分析，验证了此前报道的棉纤维起始相关生物学过程，例如细胞壁重塑。本研究还鉴定出在棉纤维起始细胞与伸长纤维中上调表达的转录因子。关键词：细胞类型比较。该基因芯片覆盖11143个基因模型：其中1个基因模型对应5种不同的寡核苷酸探针（oligonucleotide），8604个基因模型对应4种不同的寡核苷酸探针（简称oligos），1706个基因模型对应3种不同的寡核苷酸探针，831个基因模型对应3种相同的寡核苷酸探针，另有1个基因模型对应2种不同的寡核苷酸探针。每张基因芯片均与取自大田种植植株不同采收时期的1 dpa或10 dpa棉纤维RNA制备的标记核酸进行杂交，其中一组实验采用了染料互换（dye swap）标记策略。