Candidate Genes and Gene Networks Change with Age in Japanese Black Cattle by Blood Transcriptome Analysis.

Age is an important physiological factor that affects the metabolism and immune function of beef cattle. While there have been many studies using the blood transcriptome to study the effects of age on gene expression, few have been reported on beef cattle. To this end, we used the blood transcriptomes of Japanese black cattle at different ages as the study subjects and screened 1055, 345, and 1058 differential expressed genes (DEGs) in the calf vs. adult, adult vs. old, and calf vs. old comparison groups, respectively. The weighted co-expression network consisted of 1731 genes. Finally, blue, brown, and yellow age-specific modules were obtained, in which genes were enriched in signaling pathways related to growth and development and immune metabolic dysfunction, respectively. Protein-protein interaction (PPI) analysis showed gene interactions in each specific module, and 20 of the highest connectivity genes were chosen as potential hub genes. Finally, we identified 495, 244, and 1007 genes by exon-wide selection signature (EWSS) analysis of different comparison groups. Combining the results of hub genes, we found that VWF, PARVB, PRKCA, and TGFB1I1 could be used as candidate genes for growth and development stages of beef cattle. CORO2B and SDK1 could be used as candidate marker genes associated with aging. In conclusion, by comparing the blood transcriptome of calves, adult cattle, and old cattle, the candidate genes related to immunity and metabolism affected by age were identified, and the gene co-expression network of different age stages was constructed. It provides a data basis for exploring the growth, development, and aging of beef cattle.

Blood Transcriptome Analysis of Beef Cow with Different Parity Revealed Candidate Genes and Gene Networks Regulating the Postpartum Diseases.

Maternal parity is an important physiological factor influencing beef cow reproductive performance. However, there are few studies on the influence of different calving periods on early growth and postpartum diseases. Here, we conducted blood transcriptomic analysis on cows of different parities for gene discovery. We used Short Time Series Expression Miner (STEM) analysis to determine gene expression levels in cows of various parities and divided multiple parities into three main periods (nulliparous, primiparous, and multiparous) for subsequent analysis. Furthermore, the top 15,000 genes with the lowest median absolute deviation (MAD) were used to build a co-expression network using weighted correlation network analysis (WGCNA), and six independent modules were identified. Combing with Exon Wide Selection Signature (EWSS) and protein-protein interaction (PPI) analysis revealed that TPCN2, KIF22, MICAL3, RUNX2, PDE4A, TESK2, GPM6A, POLR1A, and KLHL6 involved in early growth and postpartum diseases. The GO and KEGG enrichment showed that the Parathyroid hormone synthesis, secretion, and action pathway and stem cell differentiation function-related pathways were enriched. Collectively, our study revealed candidate genes and gene networks regulating the early growth and postpartum diseases and provided new insights into the potential mechanism of reproduction advantages of different parity selection.

[Comparative study of two high-efficiency methods for purifying spermatogonial stem cells].

Objective: To compare the advantages and disadvantages of the differential attachment method and immunomagnetic bead method for purification of mouse spermatogonial stem cells (mSSCs). Methods: Ten male C57BL/6 mice aged 12～15 days were selected and sacrificed by cervical dislocation. Testes were collected and the seminiferous tubule single cell suspension was obtained by enzymatic digestion. mSSCs were purified by using the differential attachment method and immunomagnetic bead method respectively. Then a detailed comparison of the two methods in terms of cell number, separation efficiency, and impact on cell proliferation and growth was conducted. Results: Both of the methods could isolate and purify stem cells from single cell suspension of mouse seminiferous tubules. mSSCs showed typical grape cluster-like clones in vitro culture, which could be continuously cultured and proliferated for over 3 months in vitro. The testes of 10 mice could obtain 3×105±0.4×105 mSSCs (n=5) by differential attachment method, cell recovery rate (the number of cells after purification/the number of cells of the single cell suspension of seminiferous tubules) was 1.5%±0.1%; 6×105±0.4×105 mSSCs (n= 5) could be obtained by immunomagnetic bead method. The recovery rate was about 3%±0.1%, and the number of stem cells obtained by the immunomagnetic bead method was higher. The stem cells obtained by the differential attachment method were more pure, because the stem cell colonies were preferentially obtained after 5 days of in vitro culture, while the stem cells obtained by the immunomagnetic bead method needed to be cultured for about 10 days before the obvious cell colonies could be observed, but the two types of purification method had no obvious effect on the long-term growth of cells in vitro. Conclusion: Both methods can get high quality mSSCs, but both methods have their own advantages and disadvantages. The differential attachment method is more economical and practical than the other, it does not require special equipment, but the stem cell number obtained is relatively lower and the time needed is longer.