Safety Assessment Of GH-transgenic Dairy Goats

Recent years,transgenic technology has undergone rapid development,in order to improve the economic performance of animals rapidly,transgenic animals have been created continuously.However,whether the foreign gene transferred into transgenic animals bring ecological environment and human health threat,which has been debated for a long time in the scientific community and society.Thus,more comprehensive perspective safety evaluation of transgenic animals is essential and meaningful.Our precious research had produced growth hormone(GH)transgenic dairy goat by somatic cell nuclear transfer technology to improve milk production.In this study,we evaluated the own healthy,environmental safety and food safety of the GH transgenic dairy goats.Firstly,we compared the gene expression levels of non-transgenic goat(NG)and transgenic goats(TG)by high-throughput transcriptome sequencing to evaluated their own security;secondly,we collected the goat feces,intestinal contents and soil of goat farm as samples,detected the gene flow event of GH and neo,the analyzed the influence of structure of environmental bacterial by PCR-DGGE;finally,evaluated the food safety of transgenic goat milk by feeding SD rat and detected the growth speed,immunity and behavioral states of SD rat.The total experiments were divided into the following four stages:1.Gene expression changes in mammary gland between TG and NGOwn safety assessment of transgenic animal is an important part of transgenic animal safety evaluation system.It might bring unknown damage to transgenic animal when foreign gene transferred into transgenic animals.GH could improve the milk production of dairy goat through enhancing the transgenic goat mammary gland development.If this change brought health risks to transgenic goat,it would be find out by further research.In this study,we detected the gene expression in lactating dairy goat mammary gland by transcriptome sequencing for comparing the difference between TG and NG.Then,we selected 15 genes related to the development of mammary gland to confirm by qRT-PCR.The results showed that the overexpression of GH in transgenic goats could enhance the proliferation of mammary epithelial cells,but transgenic goats could maintain the proliferation on the health level by autoregulation.This results proved that GH transgenic goat could growth and develop healthily.2.Detection of foreign genes drift of TGThe environmental safety assessment is an indispensable part of the process of safety evaluation of transgenic animals,and detection of foreign genes drift is necessary in environmental safety evaluation.In this study,we detected goat feces,intestinal contents and soil of goat farm by PCR to confirm the occurrence of genes drift event.Firstly,we collected feces and intestinal contents from TG and NG.We also collected soil in goat farm as samples.Then,we extracted bacterial genome from the samples by using commercial Genome Extraction Kit.We designed the primers which targeted to the foreign genes,then detected whether the exogenous genes existed in the sample genome by PCR.The result of detection showed that there was no exogenous GH and neo in intestinal contents,feces and soil samples.This result demonstrated there was no gene drift event occurs in environment.3.Influence of the structure of the flora on the environment by GH transgenic goatWhether the species will be affected in the surrounding environment during the production of transgenic animals,it is the focus of the safety problem of transgenic animals.Evaluated the environment flora structure is an important part of environmental safety evaluation of transgenic animals.The purpose of this study was to evaluate the impact of structure of surrounding microflora by GH transgenic goat.Firstly,we amplified the V3 region segments of bacterial 16srDNA gene from the genomes of the feces,soil and intestinal contents samples.Then,separating the V3 region segments by different sequences and recovered the separated bands.Finally,sequenced the recovered DNA fragments and submitted these sequences to GenBank database,received the information of bacterial species through Blast.The results of PCR-DGGE electrophoresis similarity data analysis showed that the similarity of each kind of samples was more than 95%.The statistics of flora species of the bacteria shown that the overall distribution of bacteria in each kind sample had same dominant bacteria.This result demonstrated that GH transgenic goat did not change the structure of surrounding bacteria,and it is safety for surrounding environment.4.Food safety assessment of GH transgenic goatThe food produced by transgenic animals is also need to evaluate safety during the evaluation of safety of transgenic animals.Therefore,we evaluated the food safety of transgenic goat milk by feeding SD rat and detected the growth speed,immunity and behavioral states of SD rat.In this study,60 rats were divided into three groups and fed with GH transgenic goat milk,goat milk and normal saline maintain 90 days.During the feeding period,we observed and recorded the habits of rats,regularly weighed;collected blood samples of rats for blood and serum biochemical tests to compare the difference among groups;detected the level of IL-2,IgG,IgE and IFN-y of serum by ELISA;necropsied and weighed major organs and histopathology observation after 90 days.The results shown that there were no differences among these three groups,it demonstrated the safety of GH transgenic goat milk to a certain degree.