Scientia Agricultura Sinica ›› 2019, Vol. 52 ›› Issue (24): 4624-4631.doi: 10.3864/j.issn.0578-1752.2019.24.016

• ANIMAL SCIENCE·VETERINARY SCIENCE·RESOURCE INSECT • Previous Articles     Next Articles

Expression Pattern of KDM1A in the Development of Yak Follicles

XiangYue HUANG1,XianRong XIONG2(),Jie HAN1,XianYing YANG1,Yan WANG1,Bin WANG1,Jian LI1()   

  1. 1 College of Life Science and Technology, Southwest Minzu University, Chengdu 610041
    2 Key Laboratory of Ministry of Education for Qinghai-Tibet Plateau Animal Genetic Resources Reservation and Exploitation, Chengdu 610041
  • Received:2018-12-06 Accepted:2019-09-09 Online:2019-12-16 Published:2020-01-15
  • Contact: XianRong XIONG,Jian LI E-mail:xianrongxiong@163.com;jianli_1967@163.com

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Abstract:

【Objective】 The aim of this study was to analyze the role of the lysine-specific histone demethylase 1A (KDM1A) in follicle development and oocyte maturation of yak. 【Method】 Taking yak follicles as research objects and according to the size of the follicles, they were divided into three groups: large-sized (6.0-9.0 mm), medium-sized (3.0-5.9 mm), and small-sized (1.0-2.9 mm) follicles. And the cumulus-oocyte complexes (COCs) were collected from each group and cultured in vitro. The maturation rate of oocytes was counted and analyzed. The total RNA was extracted from oocyte and granulosa cells in each group of follicles. The real time quantitative PCR (RT-qPCR) was used to detect the relative expression of KDM1A during follicular development. The cell localization and expression of KDM1A in yak follicle were detected by immunohistochemistry, and the correlation analysis was performed by in vitro maturation and RT-qPCR. 【Result】 The maturation rate of oocytes in vitro maturation was positively related to the size of follicles and showed a rising trend with progressing of follicular development. Meanwhile, the oocyte maturation rates of large-sized, medium-sized and small-sized follicle oocytes were 90.53 %, 88.10 % and 55.14 %, respectively. The result of RT-qPCR was found that KDM1A gene of yak was widely expressed during the development of follicles, and its expression level was significantly different in developmental stages of follicles. The relative expression of mRNA in the oocytes of the large and medium-sized follicle was significantly lower than small-sized follicles (P ＜ 0.05), but the relative expression of granulosa cells in the small-sized follicle was significantly lower than large and medium-sized (P ＜ 0.01), and there was no difference in mRNA expression levels between oocytes and granulosa cells in the large and middle-sized (P ＞ 0.05). The results of immunohistochemistry showed that KDM1A was expressed in granulosa cells and membrane cells of follicles, and its expression trend was consistent with RT-qPCR. The expression of KDM1A was the highest in the large follicle and increased with the development of follicles. 【Conclusion】 The expression levels of KDM1A mRNA and protein in oocytes and granulosa cells at the development of yak follicles were dynamic, which indicated that KDM1A played an important role in follicular development and oocyte maturation. It might be related to meiosis of oocytes and proliferation and differentiation of granulosa cells, and these results of study would provide a basic data for further research of the mechanism of KDM1A in the meiosis of yak oocytes.

Key words: Yak, KDM1A, immunohistochemistry, follicle, expression

Table 1

Primer information"

基因
Gene		 引物序列(5'-3')
Sequence of primer		 退火温度
Tm(℃)		 产物长度
Product size (bp)
GAPDH F: TGCTGGTGCTGAGTATGTGGTG 60 293
R: TCTTCTGGGTGGCAGTGATGG
KDM1A F: CGCAAAGGAAACTATGTAG 60 245
R: ATTATTGAGGACGTTGAAGTC

Fig. 1

The expression of KDM1A in the oocyte (A) and the granular (B) cells of the different development stages in Yak follicle ** show extremely significant difference (P ＜ 0.01); * show significant difference (P ＜ 0.05)"

Table 1

Maturation rate of yak oocytes"

卵泡类型
Types of follicles/cm		 重复数
NO. of replicates		 卵母细胞数
NO. of COCs		 MII期卵母细胞数
NO. of COCs in MII		 成熟率
Maturation rate (%)
小 Small（1.0-2.9 mm） 3 380 209 55.14%±1.52
中 Medium（3.0-5.9 mm） 3 285 251 88.10%±0.94**
大 Large（6.0-9.0 mm） 3 169 153 90.53%±1.55**

Fig. 2

Localization of KDM1A in Yak follicle A: Localization of KDM1A in large-sized follicle of yak; B: Localization of KDM1A in medium-sized follicle of yak; C: Localization of KDM1A in small-sized follicle of yak (GC: Granular cells; TC: Theca cells)"

Fig. 3

Average density of fluorescence signals from KDM1A in Yak follicle"

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