In situ-synthesized novel microarray optimized for mouse stem cell and early developmental expression profiling
In situ-synthesized novel microarray optimized for mouse stem cell and early developmental expression profiling
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DOI：
10.1101/gr.878903
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发表时间：
2003-05-01
期刊：
影响因子：
7
通讯作者：
Ko, MSH
中科院分区：
文献类型：
--
作者：
Carter, MG;Hamatani, T;Ko, MSH
微阵列技术在小鼠胚胎学/遗传学中的应用一直受到限制，这是因为包含大量胚胎基因的微阵列不可用，以及典型的微阵列方法所需的微克量RNA与研究人员可用的极少量组织之间的差距。为了克服这些问题，我们开发了一个微阵列平台，其中包含原位合成的60聚体寡核苷酸探针，代表大约22,000个独特的小鼠转录本，主要由干细胞和胚胎cDNA文库的序列组装而成。我们已经优化了RNA标记方案和实验设计，以可靠和可重复性地使用少至2 ng的总RNA。微阵列中包含的至少98%的探针对应于我们公开可用集合中的克隆，使cDNA易于用于对感兴趣基因的进一步实验。这些特点，再加上分析非常小样本的能力，使该系统成为干细胞和胚胎基因组学研究的资源。
Applications of microarray technologies to mouse embryology/genetics have been limited, due to the nonavailability of microarrays containing large numbers of embryonic genes and the gap between microgram quantities of RNA required by typical microarray methods and the miniscule amounts of tissue available to researchers. To overcome these problems, we have developed a microarray platform containing in situ-synthesized 60-mer oligonucleotide probes representing approximately 22,000 unique mouse transcripts, assembled primarily from sequences of stem cell and embryo cDNA libraries. We have optimized RNA labeling protocols and experimental designs to use as little as 2 ng total RNA reliably and reproducibly. At least 98% of the probes contained in the microarray correspond to clones in our publicly available collections, making cDNAs readily available for further experimentation on genes of interest. These characteristics, combined with the ability to profile very small samples, make this system a resource for stem cell and embryogenomics research.