Transcriptomic Study of Spermatogenesis in the Testis of Hu Sheep and Tibetan Sheep
Abstract
:1. Introduction
2. Materials and Methods
2.1. Ethics Statement
2.2. Animals and Experimental Design
2.3. Preparation and Data Analysis of Testicular Tissue Sections
2.4. RNA Extraction, cDNA Library Preparation, and RNA-Seq
2.5. Gene Expression Quantification and Identification of Differentially Expressed Genes (DEGs)
2.6. Functional Enrichment Analysis of DEGs
2.7. RT-qPCR Analysis for the Validation of RNA-Seq Data
3. Results
3.1. Morphologic Parameters of Testis
3.2. Histological Observation of Testis
3.3. Summary of the RNA-Seq Data
3.4. Functional Categories of the Identified Genes
3.5. Analysis of Differentially Expressed Genes
3.6. GO Enrichment Analysis of DEGs
3.7. KEGG Pathway Enrichment Analysis of DEGs
3.8. Protein–Protein Interaction Network Analysis
3.9. Validation of RNA-Seq Data by RT-qPCR
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Data Availability Statement
Conflicts of Interest
References
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Breeds | Birthplace | Number | Sex | Body Weight | Age at Slaughter |
---|---|---|---|---|---|
Hu sheep | Minqin, Gansu | 3 | male | 63.23 ± 2.30 | 360 ± 5 d |
Tibetan sheep | Linxia, Gansu | 3 | male | 46.58 ± 4.17 | 360 ± 5 d |
Gene Name | Primer Sequences (5′–3′) | Product Length (bp) | Annealing Temperature (°C) |
---|---|---|---|
FOXP2 | F: GAGATTGCCCCAAACTACGAG R: GCAAATGTCCGTGTAAACCAG | 132 | 54 |
GGT5 | F: AGCTTCCTGCACAGCCCGTTC R: ACCTTCCTTGGCGATGTCCTCC | 182 | 58 |
HKDC1 | F: AGAAGAAATTACCTCTTGGCCTA R: CTCGCCTTAAACTTCTTGGTC | 134 | 49 |
IL5RA | F: GTGAATTTAACTTGCACCACA R: GACATTCTTCAATCCGAGAGC | 137 | 51 |
SH2D4B | F: CTTCCTTAACTGCAAGCCAGA R: CCATCATTGGCAGTCTACCTC | 130 | 51 |
BCL2 | F: TGAGGCTTATGAACATTCCAGT R: TTCTTCCTCCCACCCCTGCAA | 121 | 51 |
COL1A1 | F: CGCTCCTTGTGTAACTGCAT R: TTCACATGAGTCCCCATCCAC | 281 | 57 |
COL1A2 | F: GCCTATCCTTGATATTGCACCT R: CTTTTGCCCACAATTTAAGCAAG | 220 | 53 |
MMP2 | F: CATCTGGCGAACAGTGACACC R: AAGAACACAGCCTTCTCCTCC | 129 | 57 |
SOX9 | F: TGTCTAAATTCATCTGCTCCC R: TGAGCCTAAATAGACTCTGC | 271 | 56 |
β-actin | F: TGATGATCGCAGAAAGAACCC R: CTCGCTTTGAAGGTTTCCAGT | 133 | 52 |
Parameters | HS | TS |
---|---|---|
Diameter of seminiferous tubules (µm) | 152.26 ± 19.64 a | 126.35 ± 11.64 b |
Cross-sectional area of seminiferous Tubule (µm2) | 16,716.71 ± 3408.55 a | 13,482.91 ± 1780.51 b |
Seminiferous epithelium thickness (µm) | 38.82 ± 5.13 a | 34.42 ± 6.23 b |
Spermatogonia area (µm2) | 13.47 ± 2.80 a | 11.51 ± 2.572 b |
Primary spermatocyte area (µm2) | 29.27 ± 5.89 a | 25.59 ± 3.20 b |
Sperm cell diameter area (µm2) | 9.91 ± 2.25 | 8.78 ± 1.52 |
Number of spermatogonias | 37.78 ± 5.27 a | 32.08 ± 4.90 b |
Number of primary spermatocytes | 26.46 ± 4.23 a | 22.96 ± 5.27 b |
Number of spermatides | 30.02 ± 5.14 a | 24.96 ± 4.78 b |
Number of Sertoli cells | 22.83 ± 3.72 a | 15.62 ± 2.73 b |
Items | HS1 | HS2 | HS3 | TS1 | TS2 | TS3 |
---|---|---|---|---|---|---|
Raw reads | 43,732,762 | 44,297,084 | 44,120,746 | 42,299,378 | 45,277,764 | 46,281,832 |
Clean reads | 21,866,381 | 22,148,542 | 22,060,373 | 21,149,689 | 22,638,882 | 23,140,916 |
Q20 (%) | 98.42 | 98.16 | 97.48 | 98.29 | 97.41 | 97.38 |
Q30 (%) | 95.09 | 94.71 | 93.22 | 94.89 | 93.08 | 92.96 |
GCcontent (%) | 50.01 | 51.14 | 50.60 | 50.20 | 50.32 | 49.61 |
Mapped Reads (%) | 97.45 | 95.97 | 96.02 | 96.93 | 96.02 | 96.02 |
Uniq Mapped Reads (%) | 94.99 | 93.61 | 93.56 | 94.47 | 93.45 | 93.49 |
Multipl Mapped Reads (%) | 2.46 | 2.36 | 2.46 | 2.46 | 2.57 | 2.53 |
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Fu, X.; Yang, Y.; Yan, Z.; Liu, M.; Wang, X. Transcriptomic Study of Spermatogenesis in the Testis of Hu Sheep and Tibetan Sheep. Genes 2022, 13, 2212. https://doi.org/10.3390/genes13122212
Fu X, Yang Y, Yan Z, Liu M, Wang X. Transcriptomic Study of Spermatogenesis in the Testis of Hu Sheep and Tibetan Sheep. Genes. 2022; 13(12):2212. https://doi.org/10.3390/genes13122212
Chicago/Turabian StyleFu, Xiaoyu, Yanan Yang, Zunqiang Yan, Miaomiao Liu, and Xinrong Wang. 2022. "Transcriptomic Study of Spermatogenesis in the Testis of Hu Sheep and Tibetan Sheep" Genes 13, no. 12: 2212. https://doi.org/10.3390/genes13122212
APA StyleFu, X., Yang, Y., Yan, Z., Liu, M., & Wang, X. (2022). Transcriptomic Study of Spermatogenesis in the Testis of Hu Sheep and Tibetan Sheep. Genes, 13(12), 2212. https://doi.org/10.3390/genes13122212