Mao M, Fu G, Wu J S, Zhang Q H, Zhou J, Kan L X, Huang Q H, He K L, Gu B W, Han Z G, Shen Y, Gu J, Yu Y P, Xu S H, Wang Y X, Chen S J, Chen Z
Key Laboratory for Human Genome Research and Shanghai Institute of Hematology, Rui Jin Hospital, Shanghai Second Medical University, Shanghai 200025, People's Republic of China.
Proc Natl Acad Sci U S A. 1998 Jul 7;95(14):8175-80. doi: 10.1073/pnas.95.14.8175.
Hematopoietic stem/progenitor cells (HSPCs) possess the potentials of self-renewal, proliferation, and differentiation toward different lineages of blood cells. These cells not only play a primordial role in hematopoietic development but also have important clinical application. Characterization of the gene expression profile in CD34(+) HSPCs may lead to a better understanding of the regulation of normal and pathological hematopoiesis. In the present work, genes expressed in human umbilical cord blood CD34(+) cells were catalogued by partially sequencing a large amount of cDNA clones [or expressed sequence tags (ESTs)] and analyzing these sequences with the tools of bioinformatics. Among 9,866 ESTs thus obtained, 4,697 (47.6%) showed identity to known genes in the GenBank database, 2, 603 (26.4%) matched to the ESTs previously deposited in a public domain database, 1,415 (14.3%) were previously undescribed ESTs, and the remaining 1,151 (11.7%) were mitochondrial DNA, ribosomal RNA, or repetitive (Alu or L1) sequences. Integration of ESTs of known genes generated a profile including 855 genes that could be divided into different categories according to their functions. Some (8.2%) of the genes in this profile were considered related to early hematopoiesis. The possible function of ESTs corresponding to so far unknown genes were approached by means of homology and functional motif searches. Moreover, attempts were made to generate libraries enriched for full-length cDNAs, to better explore the genes in HSPCs. Nearly 60% of the cDNA clones of mRNA under 2 kb in our libraries had 5' ends upstream of the first ATG codon of the ORF. With this satisfactory result, we have developed an efficient working system that allowed fast sequencing of 32 full-length cDNAs, 16 of them being mapped to the chromosomes with radiation hybrid panels. This work may lay a basis for the further research on the molecular network of hematopoietic regulation.
造血干/祖细胞（HSPCs）具有自我更新、增殖以及向不同血细胞谱系分化的潜能。这些细胞不仅在造血发育中发挥着重要作用，还具有重要的临床应用价值。对CD34(+) HSPCs中基因表达谱的表征可能有助于更好地理解正常和病理造血的调控机制。在本研究中，通过对大量cDNA克隆[或表达序列标签（ESTs）]进行部分测序，并利用生物信息学工具分析这些序列，对人脐带血CD34(+)细胞中表达的基因进行了编目。在由此获得的9866个ESTs中，4697个（47.6%）与GenBank数据库中的已知基因具有同一性，2603个（26.4%）与先前存于公共领域数据库中的ESTs相匹配，1415个（14.3%）是先前未描述的ESTs，其余1151个（11.7%）是线粒体DNA、核糖体RNA或重复（Alu或L1）序列。已知基因的ESTs整合产生了一个包含855个基因的图谱，这些基因可根据其功能分为不同类别。该图谱中的一些基因（8.2%）被认为与早期造血有关。通过同源性和功能基序搜索探讨了对应于迄今未知基因的ESTs的可能功能。此外，还尝试构建富含全长cDNA的文库，以更好地探索HSPCs中的基因。我们文库中近60%的2 kb以下mRNA的cDNA克隆在开放阅读框（ORF）的第一个ATG密码子上游具有5'端。基于这一令人满意的结果，我们开发了一个高效的工作系统，该系统能够对32个全长cDNA进行快速测序，其中16个通过辐射杂种板定位到染色体上。这项工作可能为进一步研究造血调控的分子网络奠定基础。
