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Frontiers of Agricultural Science and Engineering

Front. Agr. Sci. Eng.    2014, Vol. 1 Issue (4) : 314-320
Effects of the TLR4 transgene on reproductive traits and DNA methylation pattern of oocytes in ewes
Yi FANG1,Xiangwei FU1,Junjie LI2,Ming DU1,Baoyu JIA1,Jinlong ZHANG3,Xiaosheng ZHANG3,Shien ZHU1,*()
1. National Engineering Laboratory for Animal Breeding and Key Laboratory of Animal Genetics, Breeding and Reproduction of Ministry of Agriculture, College of Animal Science and Technology, China Agricultural University, Beijing 100193, China
2. College of Animal Science and Technology, Agricultural University of Hebei, Baoding 071000, China
3. Animal Husbandry and Veterinary Research Institute of Tianjin, Tianjin 300412, China
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This study was conducted to systematically assess the reproductive performance of transgenic TLR4 ewes. In the TLR4 transgenic founders (F0) and their positive offspring (F1), hematological and reproductive parameters and the global DNA methylation level in oocytes at various stages were analyzed. The values of the physiological and biochemical parameters determined from the blood samples did not differ significantly between the transgenic and wild-type ewes. Moreover, the transgenic ewes showed reproductive traits similar to the wild-type ewes. These traits included characteristics of puberty, the estrus cycle, estrus duration, gestation, the pregnancy rate and the superovulation response. Additionally, no significant differences were found between transgenic and wild-type ewes in the DNA methylation level of the oocytes at various stages. In summary, the preliminary evidence presented in this paper demonstrates that the presence of the TLR4 transgene did not affect the reproductive performance in sheep.

Keywords TLR4 transgenic ewe      safety assessment      reproductive trait      oocyte      DNA methylation     
Corresponding Authors: Shien ZHU   
Online First Date: 28 January 2015    Issue Date: 10 March 2015
 Cite this article:   
Yi FANG,Xiangwei FU,Junjie LI, et al. Effects of the TLR4 transgene on reproductive traits and DNA methylation pattern of oocytes in ewes[J]. Front. Agr. Sci. Eng. , 2014, 1(4): 314-320.
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Xiangwei FU
Junjie LI
Ming DU
Baoyu JIA
Jinlong ZHANG
Xiaosheng ZHANG
Shien ZHU
Physiological Parameters F0 F1
TG (n = 5) WT (n = 5) TG (n = 7) WT (n = 6)
RBC/(1012·L-1) 12.01±0.79 12.32±0.45 12.22±0.48 12.15±0.65
WBC/(109·L-1) 18.45±3.44 20.06±3.51 17.44±2.98 20.75±4.05
HGB/(g·L-1) 112.79±8.88 107.58±5.19 110.40±4.17 110.00±12.06
HCT/% 35.15±1.28 32.77±2.48 34.34±1.15 34.95±4.15
Tab.1  Physiological parameters of TG and WT ewes in F0 and F1
Biochemical Parameters F0 F1
TG (n = 5) WT (n = 5) TG (n = 7) WT (n = 6)
TP/(g·L-1) 75.19±4.41 77.04±5.39 78.98±6.58 68.10±4.81
ALB/(g·L-1) 34.42±6.11 37.40±2.22 31.68±1.19 35.90±3.34
GLO/(g·L-1) 35.38±1.25 34.62±3.41 35.90±3.30 32.20±1.55
ALT/μL 15.29±3.29 14.38±2.77 16.67±4.12 14.50±1.52
AST/μL 151.49±10.01 139.47±7.09 148.00±12.48 133.50±7.59
BUN/(mmol·L-1) 6.66±0.42 6.01±0.29 6.60±0.23 5.85±0.35
GLU/(mmol·L-1) 3.44±0.14 3.51±0.17 3.32±0.13 3.46±0.14
TG/(mmol·L-1) 0.27±0.03 0.27±0.04 0.26±0.04 0.30±0.06
Tab.2  Biochemical parameters of TG and WT ewes in F0 and F1
Traits F0 F1
TG (n = 5) WT (n = 5) TG (n = 7) WT (n = 6)
Puberty(days) 143.91±6.14(132-151) 145.05±5.28(128-149) 145.44±6.14(131-153) 144.05±4.28(135-149)
Estrus cycle(days) 17.05±1.31(15-19) 17.87±0.36(16-18) 17.42±0.94(16-20) 17.55±0.99(16-18)
Estrus duration(hours) 33.65±2.45(24-39) 32.9±3.53(19-40) 32.65±3.31(18-41) 34.01±4.13(17-38)
Gestation(days) 148.47±1.15(145-153) 151.95±1.29(146-155) 148.41±1.67(144-152) 150.95±1.22(144-153)
Total pregnancy rate/% 100% (5/5) 100% (5/5) 100% (7/7) 100% (6/6)
Tab.3  The basic reproductive parameters of TG and WT ewes in F0 and F1
Superovulation parameters F0 F1
TG (n = 4) WT (n = 4) TG (n = 4) WT (n = 4)
Average no. of corpora lutea 12.33±2.80 15.26±1.48 16.37±3.42 14.91±1.77
Average no. of oocytes recovered 8.50±1.12 11.31±1.83 14.84±2.15 13.44±0.91
Tab.4  Superovulation response of TG and WT ewes in F0 and F1
Fig.1  Typical picture of 5-MeC patterns in TG and WT oocytes. (a)-(d) Immunofluorescence results on stage GV, GVBD, MI and MII oocytes, respectively (left, blue), PI-stained nuclei (middle), merged (right). Bar= 20 μm
Fig.2  Effect of TLR4 transgene on global DNA methylation levels in oocytes at various stages in F0 and F1. Note: The average fluorescent intensity of GV oocyte from TG group was set as 1 in (a) and (b).
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