1.Generation of genetic modified pigs devoid of GGTA1 and expressing the human leukocyte antigen-G5.
Xiaoqing ZHOU ; Yu LIU ; Chengcheng TANG ; Lingyin CHENG ; Shuwen ZHENG ; Yuling ZHENG ; Min CHEN ; Huaqiang YANG ; Qingjian ZOU ; Liangxue LAI
Chinese Journal of Biotechnology 2022;38(3):1096-1111
Pigs are considered as ideal donors for xenotransplantation because they have many physiological and anatomical characteristics similar to human beings. However, antibody-mediated immunity, which includes both natural and induced antibody responses, is a major challenge for the success of pig-to-primate xenotransplantation. Various genetic modification methods help to tailor pigs to be appropriate donors for xenotransplantation. In this study, we applied transcription activator-like effector nuclease (TALEN) to knock out the porcine α-1, 3-galactosyltransferase gene GGTA1, which encodes Gal epitopes that induce hyperacute immune rejection in pig-to-human xenotransplantation. Meanwhile, human leukocyte antigen-G5 gene HLA-G5, which acts as an immunosuppressive factor, was co-transfected with TALEN into porcine fetal fibroblasts. The cell colonies of GGTA1 biallelic knockout with positive transgene for HLA-G5 were chosen as nuclear donors to generate genetic modified piglets through a single round of somatic cell nuclear transfer. As a result, we successfully obtained 20 modified piglets that were positive for GGTA1 knockout (GTKO) and half of them expressed the HLA-G5 protein. Gal epitopes on the cell membrane of GTKO/HLA-G5 piglets were completely absent. Western blotting and immunofluorescence showed that HLA-G5 was expressed in the modified piglets. Functionally, the fibroblasts from the GTKO/HLA-G5 piglets showed enhanced resistance to complement-mediated lysis ability compared with those from GTKO-only or wild-type pigs. These results indicate that the GTKO/HLA-G5 pigs could be a valuable donor model to facilitate laboratory studies and clinics for xenotransplantation.
Animals
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Animals, Genetically Modified
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Gene Knockout Techniques
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HLA Antigens
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Humans
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Nuclear Transfer Techniques
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Swine
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Transplantation, Heterologous
2.Transgenesis for pig models.
Soo Young YUM ; Ki Young YOON ; Choong Il LEE ; Byeong Chun LEE ; Goo JANG
Journal of Veterinary Science 2016;17(3):261-268
Animal models, particularly pigs, have come to play an important role in translational biomedical research. There have been many pig models with genetically modifications via somatic cell nuclear transfer (SCNT). However, because most transgenic pigs have been produced by random integration to date, the necessity for more exact gene-mutated models using recombinase based conditional gene expression like mice has been raised. Currently, advanced genome-editing technologies enable us to generate specific gene-deleted and -inserted pig models. In the future, the development of pig models with gene editing technologies could be a valuable resource for biomedical research.
Animals
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Gene Expression
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Gene Transfer Techniques*
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Mice
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Models, Animal
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Recombinases
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Swine
3.Genetically Engineered Mouse Models for Drug Development and Preclinical Trials.
Biomolecules & Therapeutics 2014;22(4):267-274
Drug development and preclinical trials are challenging processes and more than 80% to 90% of drug candidates fail to gain approval from the United States Food and Drug Administration. Predictive and efficient tools are required to discover high quality targets and increase the probability of success in the process of new drug development. One such solution to the challenges faced in the development of new drugs and combination therapies is the use of low-cost and experimentally manageable in vivo animal models. Since the 1980's, scientists have been able to genetically modify the mouse genome by removing or replacing a specific gene, which has improved the identification and validation of target genes of interest. Now genetically engineered mouse models (GEMMs) are widely used and have proved to be a powerful tool in drug discovery processes. This review particularly covers recent fascinating technologies for drug discovery and preclinical trials, targeted transgenesis and RNAi mouse, including application and combination of inducible system. Improvements in technologies and the development of new GEMMs are expected to guide future applications of these models to drug discovery and preclinical trials.
Animals
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Drug Discovery
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Gene Transfer Techniques
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Genome
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Mice*
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Models, Animal
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United States Food and Drug Administration
4.Enhancement of ionizing radiation on liposome-mediated gene delivery in human rectal cancer HR-8348 cells.
Wenping LU ; Shiyong LI ; Ping AN ; Bo YU
Chinese Journal of Surgery 2002;40(8):604-605
OBJECTIVESTo investigate the effect of ionizing radiation on liposome-mediated gene delivery and find out a way to improve gene transfection.
METHODSPrior to liposome transfection, HR-8348 cells were irradiated at doses of 0, 2, 4, 8 Gy selected according to the surviving fraction line of HR-8348 cells after different dosage of radiation. After 36 h of liposome transfection, green fluorocytes were counted. The transfection efficiency was figured out and compared with each other.
RESULTSThe transfection efficiency of liposome-mediated gene delivery was 21.32%, 62.17%, 68.00%, 77.78% at the dose of 0, 2, 4, 8 Gy respectively and the clinical dose (2 Gy) was as high as 62.17%. Combined radiation and liposome-mediated gene delivery achieved the approximate transfection efficiency of virus vector.
CONCLUSIONIonizing radiation can improve the transfection efficiency of liposome-mediated gene delivery markedly and it is expected to treat human malignancy with liposome-mediated gene delivery combined with radiation.
Dose-Response Relationship, Radiation ; Gene Transfer Techniques ; Genetic Therapy ; Humans ; Liposomes ; Rectal Neoplasms ; therapy ; Transfection
5.Research advances in gene-enhanced tissue engineering.
Journal of Biomedical Engineering 2003;20(4):733-737
Gene-enhanced engineering deals with the scientific and technologic endeavour to produce cultured cells or polymer matrices transduced with multiple gene vectors encoding cytokine cDNA by means of genetic engineering technique, to make transduced cells or gene activated matrices highly express according cytokine, and then to enhance certain abilities of the artificial tissue. Up to now, various genes encoding modulatory species of ribonucleic of proteins such as growth factors, receptors, and transcription factors have been used in the context of gene-enhanced tissue engineering and expressed within numerous tissues, including artificial blood vessels, bone, cartilage, skin and urinary system, etc. Many experiments in vitro or in vivo have begun to show good prospects and great potential application of the new approach. We believe great changes will take place in the research field of tissue engineering due to the induction the of genetic engineering, and the new approach will become a very promising and valuable tool for therapy.
Blood Vessel Prosthesis
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Bone Substitutes
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Gene Transfer Techniques
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Genetic Engineering
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methods
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Tissue Engineering
6.Surface modification and DNA-binding assessment of nano-hydroxyapatite.
Xiong-hui WU ; Hong SUN ; Xiao-wei XING ; Li-hua HUANG ; Su-ping HUANG
Journal of Southern Medical University 2010;30(10):2233-2241
OBJECTIVETo evaluate the impact of surface modification on the DNA-binding ability of nano-hydroxyapatite (nHA).
METHODSChemical co-precipitation-hydrothermal synthesis was utilized to prepare the nHA particles, and polyethylenimine (PEI) was used for surface modification of the nHA. Transmission electron microscopic (TEM) observation and zeta potential detection of the nHA were carried out before and after surface modification. The abilities of the nanoparticles, at different pH values and different concentrations, for DNA-binding and DNA protection against nuclease digestion were assessed before and after surface modification by electrophoresis.
RESULTSTEM observation showed a short rod-like morphology of PEI-modified nHA with uniform particle size and good dispersion; the nHA without the modification tended to aggregate with poor dispersion. With a positive zeta potential, the PEI-modified nHA showed an obviously enhanced ability of DNA binding at different pH values and concentrations, with strong capacity to protect the DNA against Dnase I digestion. At the concentration of 250 µg/ml and a pH value of 7.0, the nHA-PEI showed an optimal efficiency of DNA-binding and DNA protection.
CONCLUSIONnHA with surface modification by PEI can serve as an effective vector for DNA binding and transfer.
Amino Acid Motifs ; DNA ; chemistry ; Durapatite ; chemistry ; Gene Transfer Techniques ; Genetic Vectors ; Nanoparticles ; chemistry ; Polyethyleneimine ; chemistry
7.Alzheimer's disease and nerve growth factor gene therapy.
Zhonghua HUANG ; Jian LI ; Jun ZHOU ; Jie ZHANG
Journal of Central South University(Medical Sciences) 2019;44(12):1413-1418
Gene therapy plays an important role in Alzheimer's disease (AD). In recent years, the research on gene delivery vector has gradually transferred from adenovirus vector, adeno-associated virus vector and lentivirus vector to liposomes and nanomaterial carrier systems. Graphene, the newest member of nanomaterial carrier system, has attracted extensive attention for its well permeability and biocompatibility. The methods of gene therapy can be divided into direct and indirect method. The stem cell therapy, which is the most-well studied one, belongs to the indirect method. In the gene therapy of AD, the selection of appropriate carrier and method will determine the therapeutic effect.
Alzheimer Disease
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therapy
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Dependovirus
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Gene Transfer Techniques
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Genetic Therapy
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Genetic Vectors
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Humans
8.Preparation of luciferase-expressing mRNA and expression characteristics of mRNA delivered by electroporation in vivo.
Lingjiang FAN ; Keru ZHOU ; Yanguang LIU ; Guiqin WANG ; Ting SHI ; Yihong HU ; Daixi LI
Chinese Journal of Biotechnology 2022;38(9):3379-3389
In this study, we aimed to construct a non-replication mRNA platform and explore the side effects of electroporation-mediated delivery of mRNA on the mice as well as the expression features of the mRNA. With luciferase gene as a marker, in vitro transcription with T7 RNA polymerase was carried out for the synthesis of luciferase-expressed mRNA, followed by enzymatic capping and tailing. The mRNA was delivered in vivo by electroporation via an in vivo gene delivery system, and the expression intensity and duration of luciferase in mice were observed via an in vivo imaging system. The results demonstrated that the mRNA transcripts were successfully expressed both in vitro and in vivo. The electroporation-mediated delivery of mRNA had no obvious side effects on the mice. Luciferase was expressed successfully in all the mRNA-transduced mice, while the expression intensity and duration varied among individuals. Overall, the expression level peaked on the first day after electroporation and rapidly declined on the fourth day. This study is of great importance for the construction of non-replication mRNAs and their application in vaccine or antitumor drug development.
Animals
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Electroporation/methods*
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Gene Transfer Techniques
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Luciferases/metabolism*
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Mice
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RNA, Messenger/genetics*
9.The construction of a human serum albumin small gene.
Hong-mei ZHANG ; Xiao-ling JIANG ; Huo-sheng WANG ; Li-xiong LI ; Liu-mei XU ; Ping LIN ; Mei-zhong LI ; Min WANG ; Bo-ping ZHOU
Chinese Journal of Hepatology 2006;14(6):461-462
10.Comparison of commercialization of transgenic crops in China and world-wide.
Chinese Journal of Biotechnology 2008;24(4):541-546
Currently, transgenic crops create huge economic, social and ecological benefits with the development of its commercial production. For China, the speed of development and commercialization of transgenic crops is a strategic issue for the sustainable agriculture development and the international competitiveness of our agricultural products. In this paper, we compared and analyzed the status of commercialization of transgenic crops in China and world-wide.
Agriculture
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methods
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trends
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China
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Crops, Agricultural
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genetics
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Gene Transfer Techniques
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trends
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Plants, Genetically Modified
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genetics