Large-scale Collection Of Mouse Cdna Sequences And Micro

大规模收集小鼠CDNA序列和微量

• 批准号: 6668113
• 负责人: Minoru S Ko
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6668113
• 关键词: aging; complementary DNA; developmental genetics; embryogenesis; gene expression; laboratory mouse; mammalian embryology; method development; microarray technology; nucleic acid sequence

The rationale of this project is to develop a method to monitor the expression levels of a large number of genes in various experimental conditions and to understand global changes of gene expression patterns in development and aging. The systematic analysis of expression patterns of a large number of genes is rapidly becoming the method of choice for many laboratories to understand biological systems. Accordingly, the demand for high quality cDNA libraries and cDNA microarrays is increasing. The goals of this research project are to collect mouse cDNA clones and prepare cDNA microarrays containing as many mouse genes as possible. During this period, we have assembled a 15,000 unique gene set from mouse, which consists of 2132 genes from E7.5 extraembryonic tissue cDNA library, 893 genes from E7.5 embryonic tissue cDNA library, 7692 genes from preimplantation cDNA libraries (unfertilized eggs, fertilized eggs, 2-cell, 4-cell, 8-cell, 16-cell embryos and blastocysts), and 4620 genes from E12.5 mesonephros and newborn ovary cDNA libraries. These cDNA clones were resequenced to verify clone identities and to recover more sequence information from individual cDNA clones. This "NIA Mouse 15K cDNA Clone Set" has been freely distributed to the scientific community for cDNA microarray work and other fundamental research. An additional 60,000 ESTs from various mouse organs/tissues and stem cells have also been generated and assembled into a supplementary 7.4K set, which has also been freely distributed to the research community. We have recently developed a method to isolate long-insert cDNA clones more efficiently from submicrogram quantities of total RNA. Future libraries will be made by this method. Future plans include the expansion of the set of unique genes by sequencing more cDNA clones from various embryonic collections, and the preparation and use of cDNA microarrays from the expanded set of unique genes.

这个项目的基本原理是开发一种方法来监测大量基因在各种实验条件下的表达水平，并了解基因表达模式在发育和衰老过程中的全球变化。对大量基因表达模式的系统分析正迅速成为许多实验室了解生物系统的首选方法。相应地，对高质量的cDNA库和cDNA微阵列的需求也在增加。本研究项目的目的是收集小鼠的基因克隆，并制备包含尽可能多的小鼠基因的基因芯片。在此期间，我们构建了小鼠独有的15,000个基因集，其中2132个基因来自E7.5的胚外组织cDNA文库，893个基因来自E7.5的胚胎组织文库，7692个基因来自着床前的文库(未受精卵、受精卵、2-细胞、4-细胞、8-细胞、16-细胞胚胎和囊胚)，4620个基因来自E12.5中肾和新生卵巢的cDNA文库。对这些克隆进行了重新测序，以验证克隆的同源性，并从单个克隆中恢复更多的序列信息。这一“NIA小鼠15K基因克隆集”已免费分发给科学界，用于进行基因芯片工作和其他基础研究。另外还从小鼠各种器官/组织和干细胞中提取了60,000个EST，并组装成一套补充的7.4K EST，该套EST也已免费分发给研究界。我们最近开发了一种方法，可以更有效地从亚微克级总RNA中分离出长插入的cdna克隆。未来的图书馆将用这种方法来制作。未来的计划包括通过对来自不同胚胎集合的更多cdna克隆进行测序来扩展这组独特的基因，以及从扩展的一组独特的基因中制备和使用cdna微阵列。