Abstract
Natural antisense transcripts (NATs) are endogenous transcripts that contain reverse complementary sequences to other RNAs (usually called sense transcripts). NATs regulate the expression of sense transcripts in a wide range of species. The identification and analysis of NATs are the prerequisite to elucidate their functions. Microarray is a genome-wide method to detect gene expression. However, conventional microarrays do not contain the specific probes of NATs; thus, they cannot be utilized to detect NATs. In this article, we developed a novel method to identify potential NATs with the conventional microarrays. In this method of our study, we labeled the first strand cDNA from one sample with Cy5 and labeled the second strand cDNA from another sample with Cy3, and then hybridized these labeled samples with oligonucleotide microarray. Using this method, we identified 920 potential NATs in rice variety Nipponbare. Among these potential NATs, 88 of them were confirmed by either full-length cDNA or orientated ESTs (expressed sequence tags). This is the first time that a conventional oligonucleotide microarray was employed to identify NATs in rice.
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Acknowledgments
The authors would like to thank Cindy Lim, Vuong Tran, Lama Tarayrah, and Andrew Mo for their critical readings and suggestions on the manuscript. This project was funded by grants from the Natural Science Foundation of China to LZ (3073007), GL (30670175), the National Key Basic Research Science Foundation of China to LZ (2006CB101904), GL (2007109201), and the Chinese Academy of Sciences to LZ (KSCX2-YW-N-005).
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Q. Gan and D. Li contributed equally to this work.
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12033_2011_9438_MOESM1_ESM.xls
Additional file 1: The list of 33,371 oligos that were uniquely matched to japonica known and predicted gene models. Additional file 2: The list of probes with high signal in both first and second strand cDNA-labeled samples. Additional file 3: The list of 30 potential NATs that were confirmed by genes or full-length cDNAs. Additional file 4: The list of 58 potential NATs that were confirmed by orientated ESTs. Additional file 5: The signal of oligoID_01 in first strand cDNA and oligoID_02 in second strand cDNA for the 392 oligo-pairs. The top 24 oligo-pairs have at least one oligo expressed in either the first or second strand cDNA samples. Additional file 6: The list of 12 oligos and their primers that were analyzed by RT-PCR. (XLS 6231 kb)
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Gan, Q., Li, D., Liu, G. et al. Identification of Potential Antisense Transcripts in Rice Using Conventional Microarray. Mol Biotechnol 51, 37–43 (2012). https://doi.org/10.1007/s12033-011-9438-y
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DOI: https://doi.org/10.1007/s12033-011-9438-y