Transcriptome Analysis and Identification of Differentially Expressed Transcripts of Immune-Related Genes in Spleen of Gosling and Adult Goose
Abstract
:1. Introduction
2. Results and Discussion
2.1. Histological Analysis
2.2. Illumina Sequencing and de Novo Assembly
Description | 2 Week-Gosling | Adult Goose | Both |
---|---|---|---|
RIN (RNA integrity Number) | 10.0 | 10.0 | |
Raw reads | 63,906,458 | 69,459,974 | |
Clean reads | 63,121,772 | 68,655,944 | |
Q30 (Q-score) | 93.18% | 93.25% | |
Total number of unigenes | 50,390 | ||
Total length of unigene | 79,599,367 | ||
Max length of unigene | 31,052 | ||
Min length of unigene | 301 | ||
Average length of unigene | 1579.67 | ||
N50 | 2585 |
2.3. Assembly Evaluation and Annotation
Database | Number of Annotated Unigenes | Annotated Unigene Ratio (%) |
---|---|---|
Nr | 13,841 | 27.47% |
Swiss-Prot | 11,742 | 23.30% |
GO | 9830 | 19.51% |
KOG | 9534 | 18.92% |
KEGG | 3607 | 7.16% |
unknown | 36,448 | 72.33% |
2.4. Function Classification and Pathway Analysis
2.4.1. KOG Analysis
2.4.2. GO Analysis
2.4.3. KEGG Analysis
Group Name | Unigene ID | Cytokine | Gene Description | Human | Chicken | Duck | Goose | Identity |
---|---|---|---|---|---|---|---|---|
Interleukin | CL32153Contig1 | IL6 | interleukin 6 | M54894.1 | NM_204628.1 | XM_005020542.1 | JF437643.1 | 73% |
CL35348Contig1 | IL1B | interleukin 1 beta | M15330.1 | NM_204524.1 | Absent | JF505290.1 | 93% | |
CL30075Contig1 | IL18, IL1F4 | interleukin 18 | AY044641.1 | NM_204608.1 | DQ522948.1 | JF505289.1 | 99% | |
Interferon | CL30945Contig1 | IFNA | interferon alpha | M54886.1 | EU367971.1 | KF731866.1 | HQ1155831 | 88% |
Tumor necrosis factor | CL13660Contig1 | TNFSF4, OX40L | tumor necrosis factor ligand | NM_003326.4 | XM_430147.3 | Absent | Absent | 94% |
superfamily member 4 | NM_001297562.1 | |||||||
CL33231Contig1 | TNFSF5, CD40L | tumor necrosis factor ligand | AY197739.1 | AJ293700.1 | DQ267671.2 | Absent | 99% | |
superfamily member 5 | ||||||||
CL36855Contig1 | TNFSF6, FASL | tumor necrosis factor ligand | AY225406.1 | AJ890143.1 | Absent | Absent | 99% | |
superfamily member 6 | ||||||||
CL36714Contig1 | TNFSF10, TRAIL | tumor necrosis factor ligand | NM_003810.3 NM_001190942.1 NM_001190943.1 | NM_204379.2 | XM_005017465.1 | Absent | 99% | |
superfamily member 10 | XM_005017466.1 | |||||||
CL44515Contig1 | TNFSF11, RANKL | tumor necrosis factor ligand | NM_003701.3 NM_033012.3 | NM_001083361.1 | XM_005008959.1 | Absent | 85% | |
superfamily member 11 | ||||||||
CL14930Contig1 | TNFSF15, VEGI | tumor necrosis factor ligand | NM_005118.3 NM_001204344.1 | NM_001024578.1 | XM_005022362.1 | Absent | 92% | |
superfamily member 15 | ||||||||
CL448Contig2 | TNFSF13B, BAFF | tumor necrosis factor ligand | NM_006573.4 NM_001145645.2 | NM_204327.2 | XM_005021732.1 | DQ874394.1 | 99% | |
XM_005021733.1 | ||||||||
superfamily member 13B | XM_005021734.1 | |||||||
XM_005021735.1 | ||||||||
CL34135Contig1 | EDA | ectodysplasin-A | NM_001399.4 | AY885699.1 | XM_005019557.1 | Absent | 99% | |
NM_001005609.1 | ||||||||
NM_001005612.2 | ||||||||
NM_001005610.3 | ||||||||
NM_001005613.3 | ||||||||
Transforming growth factor | CL37958Contig1 | TGFB1 | transforming growth factor beta-1 | NM_000660.5 | JQ423909.1 | Absent | Absent | 83% |
CL35089Contig1 | TGFB2 | transforming growth factor beta-2 | NM_001135599.2 NM_003238.3 | NM_001031045.3 | EU737316.1 | EF541127.1 | 99% | |
Unigene ID | Cytokine | Gene Description | Human | Chicken | Duck | Goose | Identity | |
Transforming growth factor | CL30050Contig1 | TGFB3 | transforming growth factor beta-3 | NM_003239.3 | NM_205454.1 | XM_005013912.1 | Absent | 99% |
Colony-stimulating factor | CL39251Contig1 | CSF1, MCSF | macrophage colony-stimulating factor 1 | NM_000757.5 NM_172210.2 NM_172211.3 NM_172212.2 | GQ249403.1 | XM_005015071.1 | Absent | 94% |
Chemokine | CL34468Contig1 | CCL4 | C-C motif chemokine 4 | NM_002984.3 | NM_204720.1 | Absent | Absent | 99% |
CL38070Contig1 | CCL5 | C-C motif chemokine 5 | NM_002985.2 NM_001278736.1 | NM_001045832.1 | Absent | Absent | 99% | |
CL35006Contig1 | CCL19, ELC | C-C motif chemokine 19 | NM_006274.2 | NM_001302168.1 | AY682098.1 | Absent | 100% | |
CL39853Contig1 | CX3CL1, NTT | C-X3-C motif chemokine 1 | NM_002996.4 NM_001304392.1 | NM_001077232.1 | Absent | Absent | 99% | |
CL29319Contig1 | CXCL12 | C–X–C motif chemokine 12 | NM_199168.3 NM_000609.6 | NM_204510.1 | XM_005029409.1XM_005029410.1 | Absent | 88% | |
NM_001033886.2 NM_001178134.1 | ||||||||
NM_001277990.11 | ||||||||
CL19694Contig1 | CXCL13 | C–X–C motif chemokine 13 | NM_006419.2 | FR874037.1 | Absent | Absent | 99% | |
FR874038.1 | ||||||||
CL12315Contig1 | IL8, CXCL8 | interleukin 8 | M28130.1 | HM179639.1 |
2.5. Analysis of Differentially Expressed Genes
2.6. Mining for Immunity Relevant Genes and Validation by Quantitative Real-Time PCR (qRT-PCR)
Unigene ID | Gene Symbol | Gene Description | Organism | log2 Fold Change | Pathway |
---|---|---|---|---|---|
CL22500Contig1 | IgH | immunoglobulin alpha heavy chain | Anser anser domesticus | 8.01485715881499 | HCL, NKC, BCR, FcεRI, FcγR, IIN |
CL36733Contig1 | CXCR4 | C–X–C chemokine receptor type 4 | Melopsittacus undulatus | 2.82461947963658 | CSP, LTM, IIN |
CL28326Contig1 | CXCR6 | C–X–C chemokine receptor type 6 | Anas platyrhynchos | 2.26986828363545 | CSP |
CL36371Contig1 | XCR1 | chemokine XC receptor 1 | Anas platyrhynchos | 1.9920108201249 | CSP, CCR |
CL35006Contig1 | CCL19 | chemokine C–C motif ligand 19 precursor | Anas platyrhynchos | 1.87882406908735 | CSP |
CL34468Contig1 | CCL4 | c-C motif chemokine 4-like | Anas platyrhynchos | 1.87191382409167 | CSP, TLR, CDP, CCR |
CL45402Contig1 | IgH | immunoglobulin alpha heavy chain | Anser anser domesticus | 1.86841583899162 | HCL, NKC, BCR, FcεRI, FcγR, IIN |
CL46049Contig1 | CCL26 | c-C motif chemokine 26-like | Meleagris gallopavo | 1.80369233288299 | CSP |
CL13415Contig1 | PLCG2 | 1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma-2 isoform X4 | Gallus gallus | 1.72847057141819 | NKC, BCR, FcεRI, FcγR, LTM |
CL36217Contig1 | OPN | osteopontin | Anas platyrhynchos | 1.723521984199 | TLR |
comp93267_c1_seq1 | MHCIα | MHC class I heavy chain | Anser anser | 1.55884992861107 | APP |
CL45933Contig1 | TAP2 | transporter of antigen processing 2 | Anas platyrhynchos | 1.18018757654374 | APP |
CL6275Contig1 | IL2RA | interleukin 2 receptor alpha | Anser cygnoides | 1.17080106279218 | HCL, CCR |
CL15887Contig1 | IL21R | interleukin-21 receptor isoform X1 | Anas platyrhynchos | 1.09522512352901 | CCR |
CL7617Contig1 | M3K8 | mitogen-activated protein kinase kinase kinase 8 isoform X2 | Anas platyrhynchos | 1.04305025312537 | TLR, TCR |
CL47027Contig1 | MASP1 | mannan-binding lectin serine protease 1 isoform X1 | Columba livia | – | CCC |
3.10860084300754 | |||||
CL12315Contig1 | IL8 | interleukin-8-like | Anas platyrhynchos | – | CSP, TLR, NODLR, RIGLR, CCR |
2.12963662747173 | |||||
CL36341Contig1 | CSF3R | colony stimulating factor 3 receptor (granulocyte) | Anas platyrhynchos | – | HCL |
1.92500564529962 | |||||
CL1203Contig2 | TNFRSF13B | tumor necrosis factor receptor superfamily member 13B | Anas platyrhynchos | – | IIN |
1.36191602157385 | |||||
CL20554Contig1 | ITB3 | integrin beta-3-like | Anas platyrhynchos | – | HCL |
1.34147190439098 | |||||
CL519Contig2 | TPA | tissue-type plasminogen activator | Anas platyrhynchos | – | CCC |
1.27377380687262 | |||||
CL45872Contig1 | MP2K3 | dual specificity mitogen-activated protein kinase kinase 3 | Columba livia | – | TLR, FcεRI |
1.02391521347926 |
2.6.1. Cytokines and Cytokine Receptors
2.6.2. Antigen Processing and Presentation
2.6.3. Humoral Immune Effectors
3. Materials and Methods
3.1. Ethics Statement
3.2. Sample, H&E Staining and cDNA Library Preparation
3.3. Quality Assessment and de Novo Assembly
3.4. Functional Annotation and Classification
3.5. Differential Expression Analysis and Enrichment Analysis
3.6. Identification of Candidate Genes and qRT-PCR Validation
4. Conclusions
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Wang, A.; Liu, F.; Chen, S.; Wang, M.; Jia, R.; Zhu, D.; Liu, M.; Sun, K.; Wu, Y.; Chen, X.; et al. Transcriptome Analysis and Identification of Differentially Expressed Transcripts of Immune-Related Genes in Spleen of Gosling and Adult Goose. Int. J. Mol. Sci. 2015, 16, 22904-22926. https://doi.org/10.3390/ijms160922904
Wang A, Liu F, Chen S, Wang M, Jia R, Zhu D, Liu M, Sun K, Wu Y, Chen X, et al. Transcriptome Analysis and Identification of Differentially Expressed Transcripts of Immune-Related Genes in Spleen of Gosling and Adult Goose. International Journal of Molecular Sciences. 2015; 16(9):22904-22926. https://doi.org/10.3390/ijms160922904
Chicago/Turabian StyleWang, Anqi, Fei Liu, Shun Chen, Mingshu Wang, Renyong Jia, Dekang Zhu, Mafeng Liu, Kunfeng Sun, Ying Wu, Xiaoyue Chen, and et al. 2015. "Transcriptome Analysis and Identification of Differentially Expressed Transcripts of Immune-Related Genes in Spleen of Gosling and Adult Goose" International Journal of Molecular Sciences 16, no. 9: 22904-22926. https://doi.org/10.3390/ijms160922904