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RNA-seq analysis of mRNAs and lncRNAs expression in chciken ALV-J-infected primary monocyte-derived macrophages (MDM) and uninfected control.

ABSTRACT: Purpose: The goals of this study are to investigate the differentially expressed mRNAs and lncRNAs between ALV-J infected MDM and uninfected MDM in chickens by Illumina deep sequencing. Methods: Total RNA from two ALV-J-infected MDM (designated: J3h_1, J3h_2, J36h_1 and J36h_2) and two uninfected MDM samples (designated: NC3h_1, NC3h_2, NC36h_1 and NC36h_2) was isolated by TRIzol following the manufacturer’s instruction at 3 h post infection (hpi) and 36 hpi. RNA samples were subjected to Illumina deep sequencing by Illumina Hiseq 2000. Results:Compared to the uninfected control, a total of 1568 and 550 up-regulated genes were identified in chicken MDM at 3 hpi and 36 hpi respectively, and 1227 and 397 down-regulated genes were identified at 3 hpi and 36 hpi, respectively.128 and 30 DE lncRNAs were identified in MDM at 3 hpi and 36 hpi, respectively. Conclusions: Strong immune response induced by ALV-J infection in MDM at 3 hpi. Many genes, lncRNAs involved in immune response such as PRRs signaling pathway and Jak-STAT signaling pathway at 3 hpi. Specifically, 78 ISGs expression significantly increased in ALV-J-infected MDM at 3 hpi. We speculated that host innate immune response could inhibit ALV-J replication in chicken MDM. These results provide valuable insights into the game of host antiviral immune response and ALV-J infection.

ORGANISM(S): Gallus gallus

PROVIDER: GSE103207 | GEO | 2017/08/29

SECONDARY ACCESSION(S): PRJNA400490

REPOSITORIES: GEO

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Project description:Purpose: The goals of this study are to investigate the differentially expressed miRNAs between ALV-J infected (WRR+) and uninfected (WRR-)chickens spleens by Illumina deep sequencing. Methods: 140-day-old female chickens of White Recessive Rock (WRR) were confirmed as J subgroup avian leukosis virus (ALV-J) infection. Total RNA from three ALV-J-infected spleens (designated: WRR1+, WRR2+, WRR3+) and three uninfected normal spleen samples (designated: WRR1-, WRR2-, WRR3-) was isolated by TRIzol following the manufacturer’s instruction (Invitrogen, CA, USA). RNA samples of three individuals within each group were pooled with equal amounts, and then were subjected to Illumina deep sequencing by Illumina Hiseq 2500. Results: After raw data filtered, 12,150,275 and 15,227,930 reads of 18-32 bp, representing 569,847 and 543,062 unique sequences, were obtained for WRR- and WRR+ libraries, respectively. Through blasting with the chicken reference genome, 360,180 WRR- sequences and 327,391 WRR+ sequences, which accounted for more than 60% of the unique sequences, were perfectly matched.To analyze the miRNA detection efficiency of Illimuna deep sequencing, all the clean reads were blasted with the Rfam data base 10.1, annotated and then removed rRNA, tRNA, snoRNA and other snRNAs. The annotation results revealed that miRNAs accounted for more than 68% of all clean reads in the WRR− and WRR+ libraries. In this study, a total of 476 miRNAs were identified after compared the unique sequences against the chicken miRNAs precursors in miRBase 18.0. Base on unique sequences matched counts, 167 differential expression miRNAs were identified by DEGseq package using Benjamini-q-value of 0.001 as a cut-off. In ALV-J infected spleens, 83 miRNAs showed up-regulated expression and 84 were down-regulated when compared to uninfected samples. Conclusions: Our study represents the first time to analysis of miRNA Expression in Spleen of J Subgroup Avian Leukosis Virus (ALV-J) Infected (WRR+) and Uninfected (WRR-) Broilers. A total of 167 miRNAs were found to be differentially expressed in ALV-J infected spleens when compared to uninfected chickens. These miRNAs can be considered as candidates for further study ALV-J invasion.

Project description:Purpose: The goals of this study are to investigate the differentially expressed genes between ALV-J infected (WRR+) and uninfected (WRR-)chickens spleens by Illumina deep sequencing. Methods: 140-day-old female chickens of White Recessive Rock (WRR) were confirmed as J subgroup avian leukosis virus (ALV-J) infection. Total RNA from three ALV-J-infected spleens (designated: WRR1+, WRR2+, WRR3+) and three uninfected normal spleen samples (designated: WRR1-, WRR2-, WRR3-) was isolated by TRIzol following the manufacturer’s instruction (Invitrogen, CA, USA). RNA samples of three individuals within each group were pooled with equal amounts, and then were subjected to Illumina deep sequencing by Illumina Genome Analyzer IIx. Results: Through raw data processed, 49,979,648 and 43,704,401 clean reads with an average length of 101 bp, which represented total residues of 4,859,084,087 and 4,238,826,168 bp, were obtained for WRR- and WRR+ libraries, respectively. Subsequently, the clean reads in the two libraries were assembled. Altogether, 121,493 contigs were assembled with an average length of 927 bp (ranged from 300 bp to 23,402 bp), leading to generation of 82,829 unigenes. The length of unigenes varied from 351 bp to 28,928 bp, with an average length of 1,155 bp. Based on the FPKM value of each gene, 252 DEGs were identified by DEGseq package using Benjamini-q-value of 0.05 as a cut-off. In ALV-J infected spleens, 90 genes showed up-regulated and 162 showed down-regulated expression when compared to uninfected samples. Conclusions: Our study represents the first time to elucidate the ALV-J infected chickens’spleens at the transcription level by RNA-seq technology. A total of 252 genes were found to be differentially expressed in ALV-J infected spleens when compared to uninfected chickens. These genes can be considered as candidates for further study ALV-J invasion. Spleen mRNA profiles of 140-day-old ALV-J infected (WRR+) and uninfected (WRR-) female chickens of White Recessive Rock were generated by deep sequencing, using Illumina Genome Analyzer IIx.

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RNA-seq analysis of mRNAs and lncRNAs expression in chciken ALV-J-infected primary monocyte-derived macrophages (MDM) and uninfected control.

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