Khan Shahid Y, Hackett Sean F, Lee Mei-Chong W, Pourmand Nader, Talbot C Conover, Riazuddin S Amer
The Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States.
Departpart of Biomolecular Engineering, University of California, Santa Cruz, California, United States.
Invest Ophthalmol Vis Sci. 2015 Jul;56(8):4919-26. doi: 10.1167/iovs.14-16253.
Transcriptome is the entire repertoire of transcripts present in a cell at any particular time. We undertook a next-generation whole transcriptome sequencing approach to gain insight into the transcriptional landscape of the developing mouse lens.
We ascertained mouse lenses at six developmental time points including two embryonic (E15 and E18) and four postnatal stages (P0, P3, P6, and P9). The ocular tissue at each time point was maintained as two distinct pools serving as biological replicates for each developmental stage. The mRNA and small RNA libraries were paired-end sequenced on Illumina HiSeq 2000 and subsequently analyzed using bioinformatics tools.
Mapping of mRNA and small RNA libraries generated 187.56 and 154.22 million paired-end reads, respectively. We detected a total of 14,465 genes in the mouse ocular lens at the above-mentioned six developmental stages. Of these, 46 genes exhibited a 40-fold differential (higher or lower) expression at one the five developmental stages (E18, P0, P3, P6, and P9) compared with their expression level at E15. Likewise, small RNA profiling identified 379 microRNAs (miRNAs) expressed in mouse lens at six developmental time points. Of these, 49 miRNAs manifested an 8-fold differential (higher or lower) expression at one the five developmental stages, as mentioned above compared with their expression level at E15.
We report a comprehensive profile of developing murine lens transcriptome including both mRNA and miRNA through next-generation RNA sequencing. A complete repository of the lens transcriptome of six developmental time points will be monumental in elucidating processes essential for the development of the ocular lens and maintenance of its transparency.
转录组是细胞在任何特定时间存在的所有转录本的总和。我们采用新一代全转录组测序方法，以深入了解发育中的小鼠晶状体的转录图谱。
我们确定了小鼠晶状体在六个发育时间点的情况，包括两个胚胎期（E15和E18）和四个出生后阶段（P0、P3、P6和P9）。每个时间点的眼组织被分为两个不同的样本库，作为每个发育阶段的生物学重复。mRNA和小RNA文库在Illumina HiSeq 2000上进行双末端测序，随后使用生物信息学工具进行分析。
mRNA和小RNA文库的映射分别产生了1.8756亿和1.5422亿双末端读数。我们在上述六个发育阶段的小鼠眼晶状体中总共检测到14465个基因。其中，46个基因在五个发育阶段（E18、P0、P3、P6和P9）之一与它们在E15时的表达水平相比，表现出40倍的差异（更高或更低）表达。同样，小RNA分析确定了在六个发育时间点在小鼠晶状体中表达的379种 microRNA（miRNA）。其中，49种miRNA在上述五个发育阶段之一与它们在E15时的表达水平相比，表现出8倍的差异（更高或更低）表达。
我们通过新一代RNA测序报告了发育中小鼠晶状体转录组（包括mRNA和miRNA）的全面概况。六个发育时间点的晶状体转录组的完整库对于阐明眼晶状体发育和维持其透明度所必需的过程将具有重大意义。
