Expression monitoring by hybridization to high-density oligonucleotide arrays
Expression monitoring by hybridization to high-density oligonucleotide arrays
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DOI：
10.1038/nbt1296-1675
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发表时间：
1996-12-01
影响因子：
46.9
通讯作者：
Brown, EL
中科院分区：
文献类型：
--
作者：
Lockhart, DJ;Dong, HL;Brown, EL
人类基因组编码了大约10万个不同的基因，而且几乎所有基因的至少部分序列信息将很快得到。然而，单靠序列信息不足以全面了解基因功能、表达、调控和剪接位点变异。因为细胞过程是由表达的基因库、表达的水平和时间控制的，所以有实验工具来并行地直接监测大量mrna是很重要的。我们已经开发了一种方法，是基于杂交的小，高密度阵列包含成千上万的合成寡核苷酸。该阵列仅基于序列信息设计，并使用光刻和寡核苷酸化学相结合的方法在原位合成。以1:30万的频率存在的rna被明确地检测到，并且检测是定量的超过三个数量级。这种方法为高度并行的实验研究提供了一种直接使用不断增长的序列信息的方法。由于化学的组合性质和合成包含数十万个特定寡核苷酸的小阵列的能力，该方法很容易扩展到同时监测数万个基因。
The human genome encodes approximately 100,000 different genes, and at least partial sequence information for nearly all will be available soon. Sequence information alone, however, is insufficient for a full understanding of gene function, expression, regulation, and splice-site variation. Because cellular processes are governed by the repertoire of expressed genes, and the levels and timing of expression, it is important to have experimental tools for the direct monitoring of large numbers of mRNAs in parallel. We have developed an approach that is based on hybridization to small, high-density arrays containing tens of thousands of synthetic oligonucleotides. The arrays are designed based on sequence information alone and are synthesized in situ using a combination of photolithography and oligonucleotide chemistry. RNAs present at a frequency of 1:300,000 are unambiguously detected, and detection is quantitative over more than three orders of magnitude. This approach provides a way to use directly the growing body of sequence information for highly parallel experimental investigations. Because of the combinatorial nature of the chemistry and the ability to synthesize small arrays containing hundreds of thousands of specifically chosen oligonucleotides, the method is readily scalable to the simultaneous monitoring of tens of thousands of genes.