褪黑素合成酶AANAT/ASMT基因过表达绵羊生物安全评价研究

doi:10.11843/j.issn.0366-6964.2020.07.009

Abstract

Abstract: The purpose of this experiment was to study the biological safety of sheep with melatonin synthase genes AANAT and ASMT (HIOMT) over-expressed in mammary gland. Based on the sheep models with AANAT and ASMT (HIOMT) over-expressed in mammary gland established in the previous study, we tracked the growth traits of positive transgenic sheep, analyzed the physiological and biochemical indexes of the blood and urine samples, and detected the intestinal microorganisms, melatonin levels in milk and milk composition. The results showed that:1) The body weight, body length, height and bust of positive transgenic sheep at 0, 6 and 12 months of age were not significantly different from those of ordinary sheep (P>0.05); 2) The total protein levels, albumin, globulin and cholesterol levels in blood of positive transgenic sheep as well as the number of various types of cells were not significantly different from those of ordinary sheep(P>0.05). Meantime, the levels of nitrite, protein, glucose and some other factors in urine were normal and not significantly different from those of ordinary sheep (P>0.05); 3) The sequencing results of microbiota showed that there was no significant difference in the composition of intestinal microbial communities and dominant bacteria between positive transgenic sheep and ordinary sheep(P>0.05); 4) The expression level of melatonin in blood of positive transgenic sheep was not significantly different from that of ordinary sheep (P>0.05), the level of melatonin at night was significantly higher than that at daytime (P<0.05); while the level of melatonin in milk of positive transgenic sheep was extremely significantly higher than that of ordinary sheep (P<0.01), the lactose content was significantly higher than that of ordinary sheep (P<0.05), in addition, the number of somatic cells of positive transgenic sheep was extremely significantly lower than that of ordinary sheep (P<0.01). In summary, the growth performance, many physiological and biochemical indexes and intestinal microbiota of positive transgenic sheep with AANAT and ASMT (HIOMT) over-expressed in the mammary gland were not significantly different compared with the ordinary sheep. Furthermore, the content of melatonin and lactose in milk of positive transgenic sheep were higher and the number of somatic cells was lower, which might be that melatonin in the mammary gland played an anti-inflammatory effect, reducing the number of somatic cells in the milk.

Key words: sheep, transgene, AANAT, ASMT, safety evaluation

CLC Number:

S826.2

YANG Ailing, LI Guangdong, WU Hao, JI Pengyun, ZHANG Lu, ZHANG Jinlong, ZHANG Xiaosheng, WUSIMAN Abulizi, LIAN Zhengxing, LIU Guoshi. Study of Biosafety Evaluation on Melatonin Synthase AANAT/ASMT Overexpressed Sheep[J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(7): 1563-1572.

References 47

[1]	ROBINSON J J,WIGZELL S,AITKEN R P,et al.Daily oral administration of melatonin from March onwards advances by 4 months the breeding season of ewes maintained under the ambient photoperiod at 57°N[J].Anim Reprod Sci,1992, 27(2-3):141-160.
[2]	BROWN A J,PENDERGAST J S,YAMAZAKI S.Peripheral circadian oscillators[J].Yale J Biol Med,2019,92(2):327-335.
[3]	ACUÑA-CASTROVIEJO D,ESCAMES G,VENEGAS C,et al.Extrapineal melatonin:sources,regulation,and potential functions[J]. Cell Mol Life Sci,2014,71(16):2997-3025.
[4]	DE CASTRO T B,BORDIN-JUNIOR N A,DE ALMEIDA E A,et al.Evaluation of melatonin and AFMK levels in women with breast cancer[J].Endocrine,2018,62(1):242-249.
[5]	PENG W,LEI M T,ZHANG J,et al.The protective effect of melatonin on the in vitro development of yak embryos against hydrogen peroxide-induced oxidative injury[J].Zygote,2019,27(3):118-125.
[6]	COELHO L A,ANDRADE-SILVA J,MOTTA-TEIOXEIRA L C,et al.The absence of pineal melatonin abolishes the daily rhythm of Tph1(Tryptophan Hydroxylase 1),Asmt (Acetylserotonin O-Methyltransferase),and Aanat (Aralkylamine N-Acetyltransferase) mRNA expressions in rat testes[J].Mol Neurobiol,2019,56(11):7800-7809.
[7]	YAMANAKA Y,YAMADA Y,HONMA K I,et al.Cryptochrome deficiency enhances transcription but reduces protein levels of pineal Aanat[J].J Mol Endocrinol,2018,61(4):219-229.
[8]	CAUSTON H C.Metabolic rhythms:A framework for coordinating cellular function[J].Eur J Neurosci,2020,51(1):1-12.
[9]	ARENDT J,SKENE D J.Melatonin as a chronobiotic[J].Sleep Med Rev,2005,9(1):25-39.
[10]	ZOU H J,CHEN B L,DING D,et al.Melatonin promotes the development of immature oocytes from the COH cycle into healthy offspring by protecting mitochondrial function[J].J Pineal Res,2020,68(1):e12621.
[11]	JALDO-ALBA F,MUÑOZ-HOYOS A,MOLINA-CARBALLO A,et al.Light deprivation increases plasma levels of melatonin during the first 72 h of life in human infants[J].Acta Endocrinol,1993,129(5):442-445.
[12]	MCKENNA H,REIES I K M.The case for a chronobiological approach to neonatal care[J].Early Hum Dev,2018,126:1-5.
[13]	JAN J E,WASDELL M B,FREEMAN R D,et al.Evidence supporting the use of melatonin in shortgestation infants[J].J Pineal Res,2007,42(1):22-27.
[14]	BAGHBAN R S,MOHEBBI A,VAKILZADEH G,et al.Enhancement of therapeutic DNA vaccine potency by melatonin through inhibiting VEGF expression and induction of antitumor immunity mediated by CD8+ T cells[J].Arch Virol,2018,163(3):587-597.
[15]	YANG M H,SHI J M,TIAN J H,et al.Exogenous melatonin reduces somatic cell count of milk in Holstein cows[J].Sci Rep,2017,7(1):43280.
[16]	WANG L,ZHAO Y,REITER R J,et al.Changes in melatonin levels in transgenic ‘Micro-Tom’ tomato overexpressing ovine AANAT and ovine HIOMT genes[J].J Pineal Res,2014,56(2):134-142.
[17]	MA T,TAO J L,YANG M H,et al.An AANAT/ASMT transgenic animal model constructed with CRISPR/Cas9 system serving as the mammary gland bioreactor to produce melatonin-enriched milk in sheep[J].J Pineal Res,2017,63(1):e12406.
[18]	PALMITER R D,BRINSTER R L,HAMMER R E,et al.Dramatic growth of mice that develop from eggs microinjected with metallothionein-growth hormone fusion genes[J].Nature,1982,300(5893):611-615.
[19]	UCHIDA M,SHIMATSU Y,ONOE K,et al.Production of transgenic miniature pigs by pronuclear microinjection[J].Transgenic Res,2001,10(6):577-582.
[20]	QIAN L L,TANG M X,YANG J Z,et al.Targeted mutations in myostatin by zinc-finger nucleases result in double-muscled phenotype in Meishan pigs[J].Sci Rep,2015,5(1):14435.
[21]	TIAN X Z,LV D Y,MA T,et al.AANAT transgenic sheep generated via OPS vitrified-microinjected pronuclear embryos and reproduction efficiency of the transgenic offspring[J].PeerJ,2018,6:e5420.
[22]	WANG B,BALDASSARE H,TAO T,et al.Transgenic goats produced by DNA pronuclear microinjection of in vitro derived zygotes[J].Mol Reprod Dev,2002,63(4):437-443.
[23]	WANG M,SUN Z L,ZOU Z Y,et al.Efficient targeted integration into the bovine Rosa26 locus using TALENs[J].Sci Rep,2018,8(1):10385.
[24]	COOPER D K C,GASTON R,ECKHOFF D,et al.Xenotransplantation-the current status and prospects[J].Br Med Bull,2018, 125(1):5-14.
[25]	RODRÍGUEZ-RODRÍGUEZ D R,RAMÍREZ-SOLÍS R,GARZA-ELIZONDO M A,et al.Genome editing:A perspective on the application of CRISPR/Cas9 to study human diseases (Review)[J].Int J Mol Med,2019,43(4):1559-1574.
[26]	BEHLER J,HESS W R.Approaches to study CRISPR RNA biogenesis and the key players involved[J]. Methods, 2020,172:12-26.
[27]	KALDS P,ZHOU S W,CAI B,et al.Sheep and goat genome engineering:from random transgenesis to the CRISPR era[J].Front Genet,2019,10:750.
[28]	SUN N,PETIWALA S,WANG R,et al.Development of drug-inducible CRISPR-Cas9 systems for large-scale functional screening[J]. BMC Genomics,2019,20(1):225.
[29]	KITAMOTO K,TAKETANI Y,FUJII W,et al.Gene-ration of mouse model of TGFBI-R124C corneal dystrophy using CRISPR/Cas9-mediated homology-directed repair[J].Sci Rep,2020,10(1):2000.
[30]	SUN Z L,WANG M,HAN S W,et al.Production of hypoallergenic milk from DNA-free beta-lactoglobulin (BLG) gene knockout cow using zinc-finger nucleases mRNA[J].Sci Rep,2018,8(1):15430.
[31]	WU H B,WANG Y S,ZHANG Y,et al.TALE nickase-mediated SP110 knockin endows cattle with increased resistance to tuberculosis[J].Proc Natl Acad Sci U S A,2015,112(13):E1530-E1539.
[32]	SHIGA K,UMEKI H,SHIMURA H,et al.Growth and fertility of bulls cloned from the somatic cells of an aged and infertile bull[J].Theriogenology,2005,64(2):334-343.
[33]	EDWARDS L,PEURA T,HARTWICH K,et al.Postnatal growth and circulating ACTH and cortisol concentratoins during the first month of life in cloned lambs[J].Endocrinology,2002,143(9):3699.
[34]	CHEN G S,SUI Y.Production,performance,slaughter characteristics,and meat quality of Ziwuling wild crossbred pigs[J].Trop Anim Health Prod,2018,50(2):365-372.
[35]	乔娜,干驰,朱佳琪,等.转hLF基因山羊血液生理生化指标检测分析[J].南京农业大学学报,2016,39(3):483-487.QIAO N,GAN C,ZHU J Q,et al.Hematological and biochemical indices in blood of hLF transgenic goat[J].Journal of Nanjing Agricultural University,2016,39(3):483-487.(in Chinese)
[36]	CHAVATTE-PALMER P,HEYMAN Y,RICHARD C,et al.Clinical,hormonal,and hematologic characteristics of bovine calvesderived from nuclei from somatic cells[J].Biol Reprod,2002,66(6):1596-1603.
[37]	KIM E Y,SONG D H,PARK M J,et al.Post-death cloning of endangered Jeju black cattle (Korean Native Cattle):fertility and serum chemistry in a cloned bull and cow and their offspring[J].J Reprod Dev,2013,59(6):536-543.
[38]	WANG M F,FENG S F,MA G J,et al.Whole-genome methylation analysis reveals epigenetic variation in cloned and donor pigs[J].Front Genet,2020,11:23.
[39]	XU J X,ZHAO J,WANG J W,et al.Molecular-based environmental risk assessment of three varieties of genetically engineered cows[J].Transgenic Res,2011,20(5):1043-1054.
[40]	WALSH M K,LUCEY J A,GOVINDASAMY-LUCEY S,et al.Comparison of milk produced by cows cloned by nuclear transfer with milk from non-cloned cows[J].Cloning Stem Cells,2003,5(3):213-219.
[41]	MONTAZER-TORBATI F,BOUTINAUD M,BRUN N,et al.Differences during the first lactation between cows cloned by somatic cell nuclear transfer and noncloned cows[J].J Dairy Sci,2016,99(6):4778-4794.
[42]	YONAI M,KANEYAMA K,MIYASHITA N,et al.Growth,reproduction,and lactation in somatic cell cloned cows with short telomeres[J].J Dairy Sci,2005,88(11):4097-4110.
[43]	LAIBLE G,BROPHY B,KNIGHTON D,et al.Compositional analysis of dairy products derived from clones and clonedtransgenic cattle[J].Theriogenology,2007,67(1):166-177.
[44]	TAO J L,YANG M H,WU H,et al.Effects of AANAT overexpression on the inflammatory responses and autophagy activity in the cellular and transgenic animal levels[J].Autophagy,2018,14(11):1850-1869.
[45]	TAKAHASHI M,TSUCHIYA H,HAMANO S,et al.Clinical study report on milk production in the offspring of a somatic cell cloned Holstein cow[J].J Reprod Dev,2013,59(6):595-598.
[46]	HEYMAN Y,CHAVATTE-PALMER P,BERTHELOT V,et al.Assessing the quality of products from cloned cattle:An integrative approach[J]. Theriogenology,2007,67(1):134-141.
[47]	MEHROTRA S,GOYAL V.Evaluation of designer crops for biosafety-A scientist's perspective[J].Gene,2013,515(2):241-248.

Study of Biosafety Evaluation on Melatonin Synthase AANAT/ASMT Overexpressed Sheep

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References 47

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	PENG Peiya, CHEN Yuhan, YANG Long, WANG Ming, ZHAO Ruiting, HE Jun, YIN Yulong, LIU Mei. Research Progress of Copy Number Variation in Livestock [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(4): 1356-1369.
[2]	ZHANG Shaohua, WANG Shuai, ZOU Yang, LIU Zhongli, CAI Xuepeng. Advances in Detection Approaches for Ovine Haemonchosis [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(4): 1499-1510.
[3]	YANG Yang, YU Qian, LIU Yucheng, YANG Hua, ZHAO Zhuo, WANG Limin, ZHOU Ping, YANG Qingyong, DAI Rong. Identification and Tissue Expression Analysis of the Sheep MYL Gene Family [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(4): 1551-1564.
[4]	CHANG Xindan, HU Fan, WU Zhiwu, YE Bingsen, LIU Tiehai, LIN Jie, HE Zhixiong, TAN Zhiliang. Effect of High Proportion Rumen Bypass Fat Diet on Feeding Behavior of Growing Mutton Sheep [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(3): 1077-1084.
[5]	GAO Hui, FANG Min, JIANG Lingling, MA Yaoyu, LIU Qiang, ZHANG Gang, NIU Xiaoxia, WANG Pu, LI Yong, ZHANG Sinong. Meta-analysis of Bluetongue Virus Prevalence in Sheep Flocks in China from 2012—2022 [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(2): 706-717.
[6]	WANG Haibo, ZHAN Jinshun, GU Zhiyong, CHEN Xinfeng, PAN Yue, JIA Haobin, ZHONG Xiaojun, LI Kairong, ZHAO Shengguo, HUO Junhong. Comparative Study on Meat Quality Characteristics of Three-Way Hybrid Sheep Charolais×Duper×Hu and Charolais×Australian White×Hu and Hu Sheep [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(1): 110-119.
[7]	DUAN Xiangru, KANG Jia, YANG Ruochen, SHAN Xinyu, LI Taichun, ZHAO Wen, ZHANG Yingjie, LIU Yueqin. Effect of L-cysteine on Proliferation, Apoptosis and the Secretion of Steroid Hormone in Ovine Ovarian Granulosa Cells [J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(1): 179-191.
[8]	ZHANG Hanyue, ZHAO Dan, LIANG Yu, ZHAO Bishi, FAN Mengdan, QIAO Liying, LIU Jianhua, YANG Kaijie, PAN Yangyang, LIU Wenzhong. miR-150 Regulates Ovine Preadipocyte Differentiation by Targeting AOC3 [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3262-3274.
[9]	LIU Linli, PENG Xuelan, LI Bo, CHENG Lefan, CIREN Lamu, ZHANG Enping. Effect of Overexpression of UCP3 Gene on the Differentiation of Sahu Sheep Preadipocytes [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3275-3285.
[10]	HE Mingyang, MA Yujing, WANG Yong, YANG Ruochen, LIU Yueqin, ZHANG Yingjie, DUAN Chunhui. Effects of Melatonin on Proliferation, Apoptosis of Ovarian Granulosa Cells, and Its Secretion of Steroid Hormones of Sheep [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3313-3324.
[11]	SONG Meijun, HAO Kexing, HAI Siyu, CHEN Yan, WANG Jing, HU Guangdong. Effects of SRIF-14 on Proliferation and Apoptosis of Endometrial Epithelial Cells in Sheep [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3325-3334.
[12]	LI Yuexin, LIU Aiju, MA Xiaofei, ZHENG Zhong, HU Boxin, ZHI Yunxia, TIAN Shujun. Effect of TGFβR1 on the Function of Sheep Granulosa Cells Mediated by TGF-β/Smad Signaling Pathway [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3335-3347.
[13]	MA Youji, CHEN Pengfei, MA Qing, WU Yi. Effects of Different Exercise Amounts on Rumen Microflora Diversity of Tan Sheep [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3393-3405.
[14]	LIU Xinhuan, YUN Jialei, MAO Li, LI Jizong, HAO Fei, HE Miaofeng, YANG Leilei, ZHANG Wenwen, CHENG Zilong, SUN Min, LIU Maojun, WANG Shaohui, BAI Juan, LI Wenliang. Isolation, Identification, Virulence Genes and Drug Resistance Analysis of Escherichia coli Isolated from Diarrheal Goat and Sheep [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3445-3454.
[15]	ZHANG Ying, JIN Hua, HAN Yang, SUI Dan, XIAO Xin, HAO Xiujing, LI Min. Analysis of Microbial Diversity in Nasal Cavity of Sheep with Respiratory Disease and Their Environment [J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(8): 3455-3465.