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Survey of the transcriptome of Aspergillus oryzae via massively parallel mRNA sequencing.

Wang B, Guo G, Wang C, Lin Y, Wang X, Zhao M, Guo Y, He M, Zhang Y, Pan L - Nucleic Acids Res. (2010)

Bottom Line: With the high resolution and sensitivity afforded by RNA-Seq, we were able to identify a substantial number of novel transcripts, new exons, untranslated regions, alternative upstream initiation codons and upstream open reading frames, which provide remarkable insight into the A. oryzae transcriptome.We were also able to assess the alternative mRNA isoforms in A. oryzae and found a large number of genes undergoing alternative splicing.Our analysis indicated that the transcriptome of A. oryzae is much more complex than previously anticipated, and these results may provide a blueprint for further study of the A. oryzae transcriptome.

View Article: PubMed Central - PubMed

Affiliation: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, Guangdong, China.

ABSTRACT
Aspergillus oryzae, an important filamentous fungus used in food fermentation and the enzyme industry, has been shown through genome sequencing and various other tools to have prominent features in its genomic composition. However, the functional complexity of the A. oryzae transcriptome has not yet been fully elucidated. Here, we applied direct high-throughput paired-end RNA-sequencing (RNA-Seq) to the transcriptome of A. oryzae under four different culture conditions. With the high resolution and sensitivity afforded by RNA-Seq, we were able to identify a substantial number of novel transcripts, new exons, untranslated regions, alternative upstream initiation codons and upstream open reading frames, which provide remarkable insight into the A. oryzae transcriptome. We were also able to assess the alternative mRNA isoforms in A. oryzae and found a large number of genes undergoing alternative splicing. Many genes and pathways that might be involved in higher levels of protein production in solid-state culture than in liquid culture were identified by comparing gene expression levels between different cultures. Our analysis indicated that the transcriptome of A. oryzae is much more complex than previously anticipated, and these results may provide a blueprint for further study of the A. oryzae transcriptome.

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Novel transcripts and new exons. (A) An example of transcripts previously unidentified in SC111. The green bar between the two annotated A. oryzae genes represents the novel transcript that was confirmed by RT–PCR (violet bar). (B) The length distribution of novel transcripts. (C) A new exon (blue bar) in the annotated intron of AO090003000598 was identified by RNA-Seq data.
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Figure 3: Novel transcripts and new exons. (A) An example of transcripts previously unidentified in SC111. The green bar between the two annotated A. oryzae genes represents the novel transcript that was confirmed by RT–PCR (violet bar). (B) The length distribution of novel transcripts. (C) A new exon (blue bar) in the annotated intron of AO090003000598 was identified by RNA-Seq data.

Mentions: A total of 12 074 genes, encoding proteins longer than 100 amino acids, have already been predicted using gene-finding software tools (2). Surprisingly, ∼40.75% of reads fell within the intergenic and intronic regions. These presumably were derived from as yet unrecognized transcripts and non-coding RNAs. Alternatively, they may correspond to novel exons of known genes. Extensive reads mapping and clustering revealed 1166 novel transcripts with significant expression levels above the surrounding intergenic region, of which 38.34% were longer than 500 bp (Supplementary Table S4 and Figure 3A and B). Based on the presence of ORF within the transcripts, 700 (60.03%) of the novel transcripts were likely to be protein-coding genes. To validate the novel transcripts determined by RNA-Seq, we randomly selected five of the novel transcripts and confirmed their transcriptional activity in all five cases by RT–PCR (Supplementary Figure S2). In addition to novel transcripts, 800 new candidate exons were identified in 513 annotated genes (Supplementary Table S5) by clustering successfully mapped reads, and these new exons were supported by paired-end reads. As illustrated in Figure 3C, a new candidate exon of 228 bp with average sequence depth of 11.69 was detected in the annotated gene AO090003000598, and this new candidate exon was further confirmed by RT–PCR (Supplementary Figure S2).Figure 3.


Survey of the transcriptome of Aspergillus oryzae via massively parallel mRNA sequencing.

Wang B, Guo G, Wang C, Lin Y, Wang X, Zhao M, Guo Y, He M, Zhang Y, Pan L - Nucleic Acids Res. (2010)

Novel transcripts and new exons. (A) An example of transcripts previously unidentified in SC111. The green bar between the two annotated A. oryzae genes represents the novel transcript that was confirmed by RT–PCR (violet bar). (B) The length distribution of novel transcripts. (C) A new exon (blue bar) in the annotated intron of AO090003000598 was identified by RNA-Seq data.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2926611&req=5

Figure 3: Novel transcripts and new exons. (A) An example of transcripts previously unidentified in SC111. The green bar between the two annotated A. oryzae genes represents the novel transcript that was confirmed by RT–PCR (violet bar). (B) The length distribution of novel transcripts. (C) A new exon (blue bar) in the annotated intron of AO090003000598 was identified by RNA-Seq data.
Mentions: A total of 12 074 genes, encoding proteins longer than 100 amino acids, have already been predicted using gene-finding software tools (2). Surprisingly, ∼40.75% of reads fell within the intergenic and intronic regions. These presumably were derived from as yet unrecognized transcripts and non-coding RNAs. Alternatively, they may correspond to novel exons of known genes. Extensive reads mapping and clustering revealed 1166 novel transcripts with significant expression levels above the surrounding intergenic region, of which 38.34% were longer than 500 bp (Supplementary Table S4 and Figure 3A and B). Based on the presence of ORF within the transcripts, 700 (60.03%) of the novel transcripts were likely to be protein-coding genes. To validate the novel transcripts determined by RNA-Seq, we randomly selected five of the novel transcripts and confirmed their transcriptional activity in all five cases by RT–PCR (Supplementary Figure S2). In addition to novel transcripts, 800 new candidate exons were identified in 513 annotated genes (Supplementary Table S5) by clustering successfully mapped reads, and these new exons were supported by paired-end reads. As illustrated in Figure 3C, a new candidate exon of 228 bp with average sequence depth of 11.69 was detected in the annotated gene AO090003000598, and this new candidate exon was further confirmed by RT–PCR (Supplementary Figure S2).Figure 3.

Bottom Line: With the high resolution and sensitivity afforded by RNA-Seq, we were able to identify a substantial number of novel transcripts, new exons, untranslated regions, alternative upstream initiation codons and upstream open reading frames, which provide remarkable insight into the A. oryzae transcriptome.We were also able to assess the alternative mRNA isoforms in A. oryzae and found a large number of genes undergoing alternative splicing.Our analysis indicated that the transcriptome of A. oryzae is much more complex than previously anticipated, and these results may provide a blueprint for further study of the A. oryzae transcriptome.

View Article: PubMed Central - PubMed

Affiliation: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, Guangdong, China.

ABSTRACT
Aspergillus oryzae, an important filamentous fungus used in food fermentation and the enzyme industry, has been shown through genome sequencing and various other tools to have prominent features in its genomic composition. However, the functional complexity of the A. oryzae transcriptome has not yet been fully elucidated. Here, we applied direct high-throughput paired-end RNA-sequencing (RNA-Seq) to the transcriptome of A. oryzae under four different culture conditions. With the high resolution and sensitivity afforded by RNA-Seq, we were able to identify a substantial number of novel transcripts, new exons, untranslated regions, alternative upstream initiation codons and upstream open reading frames, which provide remarkable insight into the A. oryzae transcriptome. We were also able to assess the alternative mRNA isoforms in A. oryzae and found a large number of genes undergoing alternative splicing. Many genes and pathways that might be involved in higher levels of protein production in solid-state culture than in liquid culture were identified by comparing gene expression levels between different cultures. Our analysis indicated that the transcriptome of A. oryzae is much more complex than previously anticipated, and these results may provide a blueprint for further study of the A. oryzae transcriptome.

Show MeSH