Nucleic Acids Research Advance Access published online on May 18, 2009
Nucleic Acids Research, doi:10.1093/nar/gkp334
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Genomics |
Detection of intergenic non-coding RNAs expressed in the main developmental stages in Drosophila melanogaster
1Center of Developmental Biology and Genetics, School of Life Sciences, Peking University, Beijing, China, 2Center for Bioinformatics, School of Life Sciences, Peking University, Beijing, China and 3Department of Ecology and Evolution, University of Chicago, Chicago, IL 60637, USA
*To whom correspondence should be addressed. Tel/Fax: 773 702 0577; Email: mlong{at}midway.uchicago.edu
Correspondence may also be addressed to Qichang Fan. Tel/Fax: +86 10 6275 2438; E-mail: qfan{at}pku.edu.cn
Received January 14, 2009. Revised April 20, 2009. Accepted April 21, 2009.
How many intergenically encoded non-coding RNAs (ncRNAs) are expressed during various developmental stages in Drosophila? Previous analyses in one or a few developmental stages indicated abundant expression of intergenic ncRNAs. However, some reported that ncRNAs have been recently falsified, and, in general, the false positive rate for ncRNA detection is unknown. In this report, we used reverse transcription-PCR (RT-PCR), a more robust method, to detect ncRNAs from the intergenic regions that are expressed in four major developmental stages (6–8 h embryo, 20–22 h embryo, larvae and adult). We tested 1027 regions, 
10% of all intergenic regions, and detected transcription by RT–PCR. The results from 18 342 RT–PCR experiments revealed evidence for transcription in 72.7% of intergenic regions in the developmental process. The early developmental stage appears to be associated with more abundant ncRNAs than later developmental stages. In the early stage, we detected 43.6% of intergenic regions that encode transcripts in the triplicate RT–PCR experiments, yielding an estimate of 5006 intergenic regions in the entire genome likely encoding ncRNAs. We compared the RT–PCR-related approach with previous tiling array-based approach and observed that the latter method is insensitive to short ncRNAs, especially the molecules less than 120 bp. We measured false positive rates for the analyzed genomic approaches including the RT–PCR and tiling array method.
The authors wish it to be known that, in their opinion, the first three authors should be regarded as joint First Authors.