Limits...
Analysis of differential gene expression and novel transcript units of ovine muscle transcriptomes.

Zhang C, Wang G, Wang J, Ji Z, Dong F, Chao T - PLoS ONE (2014)

Bottom Line: Among the 123,678 novel predicted transcript units (TUs), 15,015 units were predicted protein sequences.The reliability of the sequencing data was verified through qRT-PCR analysis of 12 genes.These results will provide useful information for functional genetic research in the future.

View Article: PubMed Central - PubMed

Affiliation: Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, College of Animal Science and Veterinary Medicine, Shandong Agricultural University, Taian, China ; College of Biological and Agricultural Engineering, Weifang University, Key Laboratory of Biochemistry and Molecular Biology in Universities of Shandong, Weifang, China.

ABSTRACT
In this study, we characterized differentially expressed genes (DEGs) between the muscle transcriptomes of Small-tailed Han sheep and Dorper sheep and predicted novel transcript units using high-throughput RNA sequencing technology. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed that 1,300 DEGs were involved in cellular processes, metabolic pathways, and the actin cytoskeleton pathway. Importantly, we identified 34 DEGs related to muscle cell development and differentiation. Additionally, we were able to optimize the gene structure and predict the untranslated regions (UTRs) for some of the DEGs. Among the 123,678 novel predicted transcript units (TUs), 15,015 units were predicted protein sequences. The reliability of the sequencing data was verified through qRT-PCR analysis of 12 genes. These results will provide useful information for functional genetic research in the future.

Show MeSH
qRT-PCR validation of the expressed genes using the Illumina sequencing technology.For the 12 randomly selected differentially expressed genes, fold changes of DP/SH determined from the relative Ct values of using the 2−△△CT method in qRT-PCR were compared to those detected by RPKM of DP/SH in RNA-seq. All Ct values were normalized to GAPDH and replicates (n = 3) of each sample were run.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3935930&req=5

pone-0089817-g003: qRT-PCR validation of the expressed genes using the Illumina sequencing technology.For the 12 randomly selected differentially expressed genes, fold changes of DP/SH determined from the relative Ct values of using the 2−△△CT method in qRT-PCR were compared to those detected by RPKM of DP/SH in RNA-seq. All Ct values were normalized to GAPDH and replicates (n = 3) of each sample were run.

Mentions: To confirm the reliability of the high-throughput sequencing data, we used qRT-PCR to compare the expression levels. Among 18 genes randomly selected from 1,300 DEGs, 12 genes were validated, and 6 were not detected. Of the 12 detected genes, 4 were significantly differentially expressed, whereas 8 were not in RNA-seq. The primers used for each gene and GAPDH were summarized in Table 1. Fold change of DP/SH in qRT-PCR analysis and RNA-seq were summarized in Figure 3. Of the 4 significantly differentially expressed genes, GLEAN_10009361, ENSBTAP00000018799, ENSBTAP00000041719 and ENSBTAP00000013079, fold was correspondingly big (18.0, 12.2, 18.4 and −5.0 fold, especially) in qRT-PCR. But other genes showed corresponding lower expressing fold. In addition to ENSBTAP00000018799 had a bigger difference of fold change, the results of other 11 genes were in good agreement between RNA-seq and qRT-PCR.


Analysis of differential gene expression and novel transcript units of ovine muscle transcriptomes.

Zhang C, Wang G, Wang J, Ji Z, Dong F, Chao T - PLoS ONE (2014)

qRT-PCR validation of the expressed genes using the Illumina sequencing technology.For the 12 randomly selected differentially expressed genes, fold changes of DP/SH determined from the relative Ct values of using the 2−△△CT method in qRT-PCR were compared to those detected by RPKM of DP/SH in RNA-seq. All Ct values were normalized to GAPDH and replicates (n = 3) of each sample were run.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3935930&req=5

pone-0089817-g003: qRT-PCR validation of the expressed genes using the Illumina sequencing technology.For the 12 randomly selected differentially expressed genes, fold changes of DP/SH determined from the relative Ct values of using the 2−△△CT method in qRT-PCR were compared to those detected by RPKM of DP/SH in RNA-seq. All Ct values were normalized to GAPDH and replicates (n = 3) of each sample were run.
Mentions: To confirm the reliability of the high-throughput sequencing data, we used qRT-PCR to compare the expression levels. Among 18 genes randomly selected from 1,300 DEGs, 12 genes were validated, and 6 were not detected. Of the 12 detected genes, 4 were significantly differentially expressed, whereas 8 were not in RNA-seq. The primers used for each gene and GAPDH were summarized in Table 1. Fold change of DP/SH in qRT-PCR analysis and RNA-seq were summarized in Figure 3. Of the 4 significantly differentially expressed genes, GLEAN_10009361, ENSBTAP00000018799, ENSBTAP00000041719 and ENSBTAP00000013079, fold was correspondingly big (18.0, 12.2, 18.4 and −5.0 fold, especially) in qRT-PCR. But other genes showed corresponding lower expressing fold. In addition to ENSBTAP00000018799 had a bigger difference of fold change, the results of other 11 genes were in good agreement between RNA-seq and qRT-PCR.

Bottom Line: Among the 123,678 novel predicted transcript units (TUs), 15,015 units were predicted protein sequences.The reliability of the sequencing data was verified through qRT-PCR analysis of 12 genes.These results will provide useful information for functional genetic research in the future.

View Article: PubMed Central - PubMed

Affiliation: Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, College of Animal Science and Veterinary Medicine, Shandong Agricultural University, Taian, China ; College of Biological and Agricultural Engineering, Weifang University, Key Laboratory of Biochemistry and Molecular Biology in Universities of Shandong, Weifang, China.

ABSTRACT
In this study, we characterized differentially expressed genes (DEGs) between the muscle transcriptomes of Small-tailed Han sheep and Dorper sheep and predicted novel transcript units using high-throughput RNA sequencing technology. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed that 1,300 DEGs were involved in cellular processes, metabolic pathways, and the actin cytoskeleton pathway. Importantly, we identified 34 DEGs related to muscle cell development and differentiation. Additionally, we were able to optimize the gene structure and predict the untranslated regions (UTRs) for some of the DEGs. Among the 123,678 novel predicted transcript units (TUs), 15,015 units were predicted protein sequences. The reliability of the sequencing data was verified through qRT-PCR analysis of 12 genes. These results will provide useful information for functional genetic research in the future.

Show MeSH