Construction of a high-performance human fetal liver-derived lentiviral cDNA library
Construction of a high-performance human fetal liver-derived lentiviral cDNA library
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DOI：
10.1007/s11010-008-9891-5
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发表时间：
2008-12-01
影响因子：
4.3
通讯作者：
Tani, Kenzaburo
中科院分区：
文献类型：
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作者：
Kurita, Ryo;Oikawa, Tatsuo;Tani, Kenzaburo
基因转导方法是一种非常有力的工具，不仅在基础科学，而且在临床医学。再生医学是基础医学与临床医学密切相关的一个领域。近年来，有报道称体细胞通过3 ~ 4个基因转导可产生诱导多能干细胞（iPS）。我们最近也报道了tal1/scl基因的慢病毒基因转移可以在没有基质细胞支持的情况下有效地将非人灵长类普通狨猴胚胎干细胞分化为造血细胞。在这项研究中，我们构建了一个高性能的人类胎儿肝脏来源的慢病毒表达文库，其中包含大量的个体克隆，旨在为未来的再生医学开发一个非常有用的工具，用于了解早期造血和/或肝细胞增生。我们的慢病毒cDNA文库包含超过8 × 10(7)个克隆，它们的平均插入长度为bbb20 kb。每个插入的DNA序列分析显示，bbb60 %包含许多基因的全长蛋白质编码区，包括细胞因子受体、细胞质蛋白、蛋白抑制剂和核因子。在293T细胞上的转导效率为100%，整合cDNA的平均大小约为1.1 kb。这些结果表明，我们的慢病毒人胎儿肝脏cDNA表达文库可能是一个非常有用的工具，可以加速发现参与早期造血和肝生成的新基因，并使iPS细胞更有效地应用于再生医学领域。
The gene transduction method is a very powerful tool, not only in basic science but also in clinical medicine. Regenerative medicine is one field that has close connection with both basic and clinical. Recently, it has been reported that induced pluripotent stem (iPS) cells can be produced from somatic cells by a three or four gene transduction. We have also recently reported that lentiviral gene transfer of the tal1/scl gene can efficiently differentiate non-human primate common marmoset ES cells into hematopoietic cells without the support of stromal cells. In this study, we constructed a high-performance human fetal liver-derived lentiviral expression library, which contains a high number of individual clones, in order to develop a very helpful tool for understanding early hematopoiesis and/or hepatocytosis for future regenerative medicine. Our lentiviral cDNA library consisted of more than 8 x 10(7) individual clones, and their average insert size was > 2 kb. DNA sequence analysis for each individual inserted cDNAs revealed that > 60% contained the full-length protein-coding regions for many genes including cytokine receptors, cytoplasmic proteins, protein inhibitors, and nuclear factors. The transduction efficiency on 293T cells was 100% and the average size of an integrated cDNA was similar to 1.1 kb. These results suggest that our lentiviral human fetal liver cDNA expression library could be a very helpful tool for accelerating the discovery of novel genes that are involved in early hematopoiesis and hepatopoiesis and to make the use of iPS cells more efficient in the field of regenerative medicine.