Background Intervertebral disc degeneration is the main cause of low back pain. The purpose of this study was to explore potential methods for reversing the degeneration of lumbar intervertebral discs by transplantati...Background Intervertebral disc degeneration is the main cause of low back pain. The purpose of this study was to explore potential methods for reversing the degeneration of lumbar intervertebral discs by transplantation of gene-modified nucleus pulposus cells into rabbit degenerative lumbar intervertebral discs after transfecting rabbit nucleus pulposus cells with adeno-associated virus 2 （AAV2）-mediated connective tissue growth factor （CTGF） and tissue inhibitor of metalloproteinases 1 （TIMP1） genes in vitro. Methods Computer tomography （CT）-guided percutaneous annulus fibrosus injury was performed to build degenerative lumbar intervertebral disc models in 60 New Zealand white rabbits, rAAV2-CTGF-IRES-TIMPI-transfected rabbit nucleus pulposus cells were transplanted into degenerative lumbar intervertebral discs （transplantation group）, phosphate-buffered saline （PBS） was injected into degenerative lumbar intervertebral discs （degeneration control group） and normal lumbar intervertebral discs served as a blank control group. After 6, 10 and 14 weeks, the disc height index （DHI） and signal intensity in intervertebral discs were observed by X-ray and magnetic resonance imaging （MRI） analysis The expression of CTGF and TIMP1 in nucleus pulposus tissue was determined by Western blotting analysis, the synthesis efficiency of proteoglycan was determined by a 35S-sulfate incorporation assay, and the mRNA expression of type II collagen and proteoglycan was detected by RT-PCR. Results MRI confirmed that degenerative intervertebral discs appeared two weeks after percutaneous puncture. Transgenic nucleus pulposus cell transplantation could retard the rapid deterioration of the DHI. MRI indicated that degenerative intervertebral discs were relieved in the transplantation group compared with the degeneration control group. The expression of collagen II mRNA and proteoglycan mRNA was significantly higher in the transplantation group and the blank control group compared with the degeneration 展开更多
Targeting of the synthesized polypeptide in the cells is an important research field in modern cell biology. Cowpea trypsin inhibitor (cpti) gene has been modified and a fusion protein gene (sck) was produced by fusin...Targeting of the synthesized polypeptide in the cells is an important research field in modern cell biology. Cowpea trypsin inhibitor (cpti) gene has been modified and a fusion protein gene (sck) was produced by fusing a signal peptide sequence at cpti 5' end and an endoplasm reticulum (ER) retention signal peptide at cpti3' end respectively. The signal peptide can direct the newly synthesized polypeptide into ER, while ER retention signal can make the protein retained in the ER and its derivative protein body. ELISA test indicated that the accumulation level of foreign CpTI protein in sck transgenic tobacco (Nicotiana tabacum L.) was two times higher than cpti transgenic tobaccos and some individuals were four times higher. At the same time, sck transgenic tobacco has a high resistance to Lepidoptera pest due to the increased accumulation level of foreign CpTI protein. The strategy of foreign protein targeting can be used to increase the accumulation level of foreign protein in transgenic plants and can be widely applied to other related research field in plant genetic engineering.展开更多
Cotton plays a crucial role in shaping Indian economy and rural livelihoods.The cotton crop is prone to numerous insect pests,necessitating insecticidal application,which increases production costs.The advent of the e...Cotton plays a crucial role in shaping Indian economy and rural livelihoods.The cotton crop is prone to numerous insect pests,necessitating insecticidal application,which increases production costs.The advent of the expression of Bacillus thuringiensis(Bt)insecticidal protein in cotton has significantly reduced the burden of pest without compromising environmental or human health.After the introduction of transgenic cotton,the cultivated area expanded to 22 million hectares,with a 64% increase in adoption by farmers worldwide.Currently,Bt cotton accounts for 93% of the cultivated cotton area in India.However,extensive use of Bt cotton has accelerated resistance development in pests like the pink bollworm.Furthermore,the overreliance on Bt cotton has reduced the use of broad-spectrum pesticides,favouring the emergence of secondary pests with significant challenges.This emphasizes the urgent necessity for developing novel pest management strategies.The high-dose and refuge strategy was initially effective for managing pest resistance in Bt cotton,but its implementation in India faced challenges due to misunderstandings about the use of non-Bt refuge crops.Although gene pyramiding was introduced as a solution,combining mono toxin also led to instances of cross-resistance.Therefore,there is a need for further exploration of biotechnological approaches to manage insect resistance in Bt cotton.Advanced biotechnological strategies,such as sterile insect release,RNA interference(RNAi)-mediated gene silencing,stacking Bt with RNAi,and genome editing using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein(CRISPR-Cas),offer promising tools for identifying and managing resistance genes in insects.Additionally,CRISPR-mediated gene drives and the development of novel biopesticides present potential avenues for effective pest management in cotton cultivation.These innovative approaches could significantly enhance the sustainability and efficacy of pest resistance management in Bt cotton.展开更多
[Objective] The aim was to construct a plasmid reference molecule (PRM) for detection of transgenic soybean MON89788. [Method] the lectin gene sequence,3'-junction and 5'-junction sequence between host plant D...[Objective] The aim was to construct a plasmid reference molecule (PRM) for detection of transgenic soybean MON89788. [Method] the lectin gene sequence,3'-junction and 5'-junction sequence between host plant DNA integrated DNA of MON89788 soybean were amplified independently,and the three fragments were cloned into the cloning vector pMD18-T in order through molecular manipulation method to construct pMD-LM3M5,the applicability of the constructed novel PRM was tested. [Result] Sequencing confirmation result showed that the PRM was 3 700 bp in length,containing 1 029 bp of recombined DNA fragment. The limits of qualitative detection of the PRM were 10 copies. [Conclusion] The PRM constructed in this study was suitable for the identification of MON89788 event.展开更多
Some 20 years ago,the EU introduced complex regulatory rules for the growth of transgenic crops,which resulted in a de facto ban to grow these plants in fields within most European countries.With the rise of novel gen...Some 20 years ago,the EU introduced complex regulatory rules for the growth of transgenic crops,which resulted in a de facto ban to grow these plants in fields within most European countries.With the rise of novel genome editing technologies,it has become possible to improve crops genetically in a directed way without the need for incorporation of foreign genes.Unfortunately,in 2018,the European Court of Justice ruled that such gene-edited plants are to be regulated like transgenic plants.Since then,European scientists and breeders have challenged this decision and requested a revision of this outdated law.Finally,after 5 years,the European Commission has now published a proposal on how,in the future,to regulate crops produced by new breeding technologies.The proposal tries to find a balance between the different interest groups in Europe.On one side,genetically modified plants,which cannot be discerned from their natural counterparts,will exclusively be used for food and feed and are-besides a registration step-not to be regulated at all.On the other side,plants expressing herbicide resistance are to be excluded from this regulation,a concession to the strong environmental associations and NGOs in Europe.Moreover,edited crops are to be excluded from organic farming to protect the business interests of the strong organic sector in Europe.Nevertheless,if this law passes European parliament and council,unchanged,it will present a big step forward toward establishing a more sustainable European agricultural system.Thus,it might soon be possible to develop and grow crops that are more adapted to global warming and whose cultivation will require lower amounts of pesticides.However,there is still a long way to go until the law is passed.Too often,the storm of arguments raised by the opponents,based on irrational fears of mutations and a naive understanding of nature,has fallen on fruitful ground in Europe.展开更多
Functional control of CRISPR/Cas9 is essential for precise gene manipulation.Chemical engineering of guide RNA(gRNA)provides diverse approaches for conditional control of CRISPR/Cas9 function with a variety of chemica...Functional control of CRISPR/Cas9 is essential for precise gene manipulation.Chemical engineering of guide RNA(gRNA)provides diverse approaches for conditional control of CRISPR/Cas9 function with a variety of chemical reactive groups.However,previous investigations into chemically engineering gRNA only unidirectionally regulated the CRISPR/Cas9 function via stimuli-induced caging/decaging processes.Herein,we propose a combinatory strategy to engineer the dynamics of gRNA in which photocontrolled strand-displacement reactions coupled with sequence designs of gRNA can achieve lightinduced switching-on/off control of CRISPR/Cas9 function.Biochemical analysis and cellular gene regulation indicate this approach is capable of both activating and deactivating CRISPR/Cas9 activities using light irradiation.Moreover,photocontrolled multiplex modulations of gene expression for opposite regulatory effects have also been achieved simultaneously under the same cellular context.This work establishes an essential principle for construction of stimuli-induced switching-on/off modulations of gRNA that can greatly enrich the versatility of conditional control for a variety of CRISPR/Cas9-based applications.展开更多
To ensure safe use of genetically modified organisms(GMOs),since 1993,China has made great efforts to establish and improve the safety regulatory system for GMOs.Here,we summarize and analyze the regulatory framework ...To ensure safe use of genetically modified organisms(GMOs),since 1993,China has made great efforts to establish and improve the safety regulatory system for GMOs.Here,we summarize and analyze the regulatory framework of agricultural GMOs,and the progress in regulatory approval of GM crops in China.In general,the development of GMO safety regulations underwent four stages:exploration(1993–2000),development(2001–2010),improvement(2011–2020)and current(2021-present)stage.The first formal regulation was promulgated in 1993,which provided a basis for further development of the regulations,during the exploration stage,when insect-resistant GM cotton,expressing genes from Bacillus thuringiensis(Bt),was approved for cultivation.During the development stage,the Chinese government issued a series of administrative measures,which covered almost all the fields relative to GMO safety when the basic regulatory system was established.Along with the controversy over GMO safety,the regulations have been further,and greatly improved,during improvement stage.From 2021,a few additional revisions have been made,and meanwhile,the new regulation on gene-edited crops was introduced with the development of biotechnology,forming a relative complete regulation and law system for China.The well-developed GMO regulations establishes a firm basis for safe use of GM crops in China.Currently,GM cotton and GM papaya have been widely grown on a large scale in China that have brought great economic and ecological benefits.In addition,12 corn events,3 soybean events,and 2 rice events have also obtained biosafety certification,but presently,these lines have yet to enter commercial production.However,several GM soybean and corn events have entered pilot industrialization,and can soon be expected to be commercially grown in China.In addition to planting,six GM crops,including soybean,corn,cotton,canola,papaya and sugar beet,with a total of 64 events,have been approved for import as processing material in China.展开更多
Recombinant adeno-associated virus （rAAV） vectors have been extensively used for experimental gene therapy of inherited human diseases. Several advantages, such as simple vector construction, high targeting frequenc...Recombinant adeno-associated virus （rAAV） vectors have been extensively used for experimental gene therapy of inherited human diseases. Several advantages, such as simple vector construction, high targeting frequency by homologous recombination, and applica- bility to many cell types, make rAAV an attractive approach for targeted genome editing. Combined with cloning by somatic cell nuclear transfer （SCNT）, this technology has recently been successfully adapted to generate gene-targeted pigs as models for cystic fibrosis, hereditary tyrosinemia type 1, and breast cancer. This review summarizes the development of rAAV for targeted genome editing in mammalian cells and provides strategies for enhancing the rAAV-mediated targeting frequency by homologous recombination. We discuss current development and application of the rAAV vectors for targeted genome editing in porcine primary fibroblasts, which are subse- quently used as donor cells for SCNT to generate cloned genetically designed pigs and provide positive perspectives for the generation of gene-targeted pigs with rAAV in the future.展开更多
We previously demonstrated that gene-modified umbilical cord blood mononuclear cells overexpressing a combination of recombinant neurotrophic factors are a promising therapeutic approach for cell-mediated gene therapy...We previously demonstrated that gene-modified umbilical cord blood mononuclear cells overexpressing a combination of recombinant neurotrophic factors are a promising therapeutic approach for cell-mediated gene therapy for neurodegenerative diseases,neurotrauma,and stroke.In this study,using a mini pig model of spinal cord injury,we proposed for the first time the use of gene-modified leucoconcentrate prepared from peripheral blood in the plastic blood bag for personalized ex vivo gene therapy.Leucoconcentrate obtained from mini pig peripheral blood was transduced with a chimeric adenoviral vector(Ad5/35 F)that carried an enhanced green fluorescent protein(EGFP)reporter gene in the plastic blood bag.The day after blood donation,the mini pigs were subjected to moderate SCI and four hours post-surgery they were intravenously autoinfused with gene-modified leucoconcentrate.A week after gene-modified leucoconcentrate therapy,fluorescent microscopy revealed EGFP-expressing leucocytes in spinal cord at the site of contusion injury.In the spleen the groups of EGFP-positive cells located in the lymphoid follicles were observed.In vitro flow cytometry and fluorescent microscopy studies of the gene-modified leucoconcentrate samples also confirmed the production of EGFP by leucocytes.Thus,the efficacy of leucocytes transduction in the plastic blood bag and their migratory potential suggest their use for temporary production of recombinant biologically active molecules to correct certain pathological conditions.This paper presents a proof-of-concept of simple,safe and effective approach for personalized ex vivo gene therapy based on gene-modified leucoconcentrate autoinfusion.The animal protocols were approved by the Kazan State Medical University Animal Care and Use Committee(approval No.5)on May 27,2014.展开更多
基金This study was supported by a grant from the National Natural Science Foundation of China （No. 30471750）.
文摘Background Intervertebral disc degeneration is the main cause of low back pain. The purpose of this study was to explore potential methods for reversing the degeneration of lumbar intervertebral discs by transplantation of gene-modified nucleus pulposus cells into rabbit degenerative lumbar intervertebral discs after transfecting rabbit nucleus pulposus cells with adeno-associated virus 2 （AAV2）-mediated connective tissue growth factor （CTGF） and tissue inhibitor of metalloproteinases 1 （TIMP1） genes in vitro. Methods Computer tomography （CT）-guided percutaneous annulus fibrosus injury was performed to build degenerative lumbar intervertebral disc models in 60 New Zealand white rabbits, rAAV2-CTGF-IRES-TIMPI-transfected rabbit nucleus pulposus cells were transplanted into degenerative lumbar intervertebral discs （transplantation group）, phosphate-buffered saline （PBS） was injected into degenerative lumbar intervertebral discs （degeneration control group） and normal lumbar intervertebral discs served as a blank control group. After 6, 10 and 14 weeks, the disc height index （DHI） and signal intensity in intervertebral discs were observed by X-ray and magnetic resonance imaging （MRI） analysis The expression of CTGF and TIMP1 in nucleus pulposus tissue was determined by Western blotting analysis, the synthesis efficiency of proteoglycan was determined by a 35S-sulfate incorporation assay, and the mRNA expression of type II collagen and proteoglycan was detected by RT-PCR. Results MRI confirmed that degenerative intervertebral discs appeared two weeks after percutaneous puncture. Transgenic nucleus pulposus cell transplantation could retard the rapid deterioration of the DHI. MRI indicated that degenerative intervertebral discs were relieved in the transplantation group compared with the degeneration control group. The expression of collagen II mRNA and proteoglycan mRNA was significantly higher in the transplantation group and the blank control group compared with the degeneration
文摘Targeting of the synthesized polypeptide in the cells is an important research field in modern cell biology. Cowpea trypsin inhibitor (cpti) gene has been modified and a fusion protein gene (sck) was produced by fusing a signal peptide sequence at cpti 5' end and an endoplasm reticulum (ER) retention signal peptide at cpti3' end respectively. The signal peptide can direct the newly synthesized polypeptide into ER, while ER retention signal can make the protein retained in the ER and its derivative protein body. ELISA test indicated that the accumulation level of foreign CpTI protein in sck transgenic tobacco (Nicotiana tabacum L.) was two times higher than cpti transgenic tobaccos and some individuals were four times higher. At the same time, sck transgenic tobacco has a high resistance to Lepidoptera pest due to the increased accumulation level of foreign CpTI protein. The strategy of foreign protein targeting can be used to increase the accumulation level of foreign protein in transgenic plants and can be widely applied to other related research field in plant genetic engineering.
文摘Cotton plays a crucial role in shaping Indian economy and rural livelihoods.The cotton crop is prone to numerous insect pests,necessitating insecticidal application,which increases production costs.The advent of the expression of Bacillus thuringiensis(Bt)insecticidal protein in cotton has significantly reduced the burden of pest without compromising environmental or human health.After the introduction of transgenic cotton,the cultivated area expanded to 22 million hectares,with a 64% increase in adoption by farmers worldwide.Currently,Bt cotton accounts for 93% of the cultivated cotton area in India.However,extensive use of Bt cotton has accelerated resistance development in pests like the pink bollworm.Furthermore,the overreliance on Bt cotton has reduced the use of broad-spectrum pesticides,favouring the emergence of secondary pests with significant challenges.This emphasizes the urgent necessity for developing novel pest management strategies.The high-dose and refuge strategy was initially effective for managing pest resistance in Bt cotton,but its implementation in India faced challenges due to misunderstandings about the use of non-Bt refuge crops.Although gene pyramiding was introduced as a solution,combining mono toxin also led to instances of cross-resistance.Therefore,there is a need for further exploration of biotechnological approaches to manage insect resistance in Bt cotton.Advanced biotechnological strategies,such as sterile insect release,RNA interference(RNAi)-mediated gene silencing,stacking Bt with RNAi,and genome editing using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein(CRISPR-Cas),offer promising tools for identifying and managing resistance genes in insects.Additionally,CRISPR-mediated gene drives and the development of novel biopesticides present potential avenues for effective pest management in cotton cultivation.These innovative approaches could significantly enhance the sustainability and efficacy of pest resistance management in Bt cotton.
基金Supported by Major Projects of Cultivating New Varieties by Trans-genic Technology (2008ZX08012-001)~~
文摘[Objective] The aim was to construct a plasmid reference molecule (PRM) for detection of transgenic soybean MON89788. [Method] the lectin gene sequence,3'-junction and 5'-junction sequence between host plant DNA integrated DNA of MON89788 soybean were amplified independently,and the three fragments were cloned into the cloning vector pMD18-T in order through molecular manipulation method to construct pMD-LM3M5,the applicability of the constructed novel PRM was tested. [Result] Sequencing confirmation result showed that the PRM was 3 700 bp in length,containing 1 029 bp of recombined DNA fragment. The limits of qualitative detection of the PRM were 10 copies. [Conclusion] The PRM constructed in this study was suitable for the identification of MON89788 event.
文摘Some 20 years ago,the EU introduced complex regulatory rules for the growth of transgenic crops,which resulted in a de facto ban to grow these plants in fields within most European countries.With the rise of novel genome editing technologies,it has become possible to improve crops genetically in a directed way without the need for incorporation of foreign genes.Unfortunately,in 2018,the European Court of Justice ruled that such gene-edited plants are to be regulated like transgenic plants.Since then,European scientists and breeders have challenged this decision and requested a revision of this outdated law.Finally,after 5 years,the European Commission has now published a proposal on how,in the future,to regulate crops produced by new breeding technologies.The proposal tries to find a balance between the different interest groups in Europe.On one side,genetically modified plants,which cannot be discerned from their natural counterparts,will exclusively be used for food and feed and are-besides a registration step-not to be regulated at all.On the other side,plants expressing herbicide resistance are to be excluded from this regulation,a concession to the strong environmental associations and NGOs in Europe.Moreover,edited crops are to be excluded from organic farming to protect the business interests of the strong organic sector in Europe.Nevertheless,if this law passes European parliament and council,unchanged,it will present a big step forward toward establishing a more sustainable European agricultural system.Thus,it might soon be possible to develop and grow crops that are more adapted to global warming and whose cultivation will require lower amounts of pesticides.However,there is still a long way to go until the law is passed.Too often,the storm of arguments raised by the opponents,based on irrational fears of mutations and a naive understanding of nature,has fallen on fruitful ground in Europe.
基金the National Natural Science Foundation of China(grant nos.21977122 and 22222706)the National Key R&D Program of China(grant no.2020YFA0211200)the Guangdong Basic Research Center of Excellence for Functional Molecular Engineering.
文摘Functional control of CRISPR/Cas9 is essential for precise gene manipulation.Chemical engineering of guide RNA(gRNA)provides diverse approaches for conditional control of CRISPR/Cas9 function with a variety of chemical reactive groups.However,previous investigations into chemically engineering gRNA only unidirectionally regulated the CRISPR/Cas9 function via stimuli-induced caging/decaging processes.Herein,we propose a combinatory strategy to engineer the dynamics of gRNA in which photocontrolled strand-displacement reactions coupled with sequence designs of gRNA can achieve lightinduced switching-on/off control of CRISPR/Cas9 function.Biochemical analysis and cellular gene regulation indicate this approach is capable of both activating and deactivating CRISPR/Cas9 activities using light irradiation.Moreover,photocontrolled multiplex modulations of gene expression for opposite regulatory effects have also been achieved simultaneously under the same cellular context.This work establishes an essential principle for construction of stimuli-induced switching-on/off modulations of gRNA that can greatly enrich the versatility of conditional control for a variety of CRISPR/Cas9-based applications.
基金supported by the National Special Biological Breeding Program of the People’s Republic of China.
文摘To ensure safe use of genetically modified organisms(GMOs),since 1993,China has made great efforts to establish and improve the safety regulatory system for GMOs.Here,we summarize and analyze the regulatory framework of agricultural GMOs,and the progress in regulatory approval of GM crops in China.In general,the development of GMO safety regulations underwent four stages:exploration(1993–2000),development(2001–2010),improvement(2011–2020)and current(2021-present)stage.The first formal regulation was promulgated in 1993,which provided a basis for further development of the regulations,during the exploration stage,when insect-resistant GM cotton,expressing genes from Bacillus thuringiensis(Bt),was approved for cultivation.During the development stage,the Chinese government issued a series of administrative measures,which covered almost all the fields relative to GMO safety when the basic regulatory system was established.Along with the controversy over GMO safety,the regulations have been further,and greatly improved,during improvement stage.From 2021,a few additional revisions have been made,and meanwhile,the new regulation on gene-edited crops was introduced with the development of biotechnology,forming a relative complete regulation and law system for China.The well-developed GMO regulations establishes a firm basis for safe use of GM crops in China.Currently,GM cotton and GM papaya have been widely grown on a large scale in China that have brought great economic and ecological benefits.In addition,12 corn events,3 soybean events,and 2 rice events have also obtained biosafety certification,but presently,these lines have yet to enter commercial production.However,several GM soybean and corn events have entered pilot industrialization,and can soon be expected to be commercially grown in China.In addition to planting,six GM crops,including soybean,corn,cotton,canola,papaya and sugar beet,with a total of 64 events,have been approved for import as processing material in China.
基金supported by the grants from the Danish National Researeh Infrastructure Programme to the Danish Genetieally Modified Animal Resource(DAG- MAR)as well as from the"Sino一Danish Breast Caneer Research Centre"under the ausPiees of the Danish National Researeh Foundation(Grundforskningsfonden)the National Natural Seience Foundation of China
文摘Recombinant adeno-associated virus （rAAV） vectors have been extensively used for experimental gene therapy of inherited human diseases. Several advantages, such as simple vector construction, high targeting frequency by homologous recombination, and applica- bility to many cell types, make rAAV an attractive approach for targeted genome editing. Combined with cloning by somatic cell nuclear transfer （SCNT）, this technology has recently been successfully adapted to generate gene-targeted pigs as models for cystic fibrosis, hereditary tyrosinemia type 1, and breast cancer. This review summarizes the development of rAAV for targeted genome editing in mammalian cells and provides strategies for enhancing the rAAV-mediated targeting frequency by homologous recombination. We discuss current development and application of the rAAV vectors for targeted genome editing in porcine primary fibroblasts, which are subse- quently used as donor cells for SCNT to generate cloned genetically designed pigs and provide positive perspectives for the generation of gene-targeted pigs with rAAV in the future.
基金the Russian Science Foundation(No.16-15-00010to RRI)the Russian Government Program of Competitive Growth of Kazan Federal University。
文摘We previously demonstrated that gene-modified umbilical cord blood mononuclear cells overexpressing a combination of recombinant neurotrophic factors are a promising therapeutic approach for cell-mediated gene therapy for neurodegenerative diseases,neurotrauma,and stroke.In this study,using a mini pig model of spinal cord injury,we proposed for the first time the use of gene-modified leucoconcentrate prepared from peripheral blood in the plastic blood bag for personalized ex vivo gene therapy.Leucoconcentrate obtained from mini pig peripheral blood was transduced with a chimeric adenoviral vector(Ad5/35 F)that carried an enhanced green fluorescent protein(EGFP)reporter gene in the plastic blood bag.The day after blood donation,the mini pigs were subjected to moderate SCI and four hours post-surgery they were intravenously autoinfused with gene-modified leucoconcentrate.A week after gene-modified leucoconcentrate therapy,fluorescent microscopy revealed EGFP-expressing leucocytes in spinal cord at the site of contusion injury.In the spleen the groups of EGFP-positive cells located in the lymphoid follicles were observed.In vitro flow cytometry and fluorescent microscopy studies of the gene-modified leucoconcentrate samples also confirmed the production of EGFP by leucocytes.Thus,the efficacy of leucocytes transduction in the plastic blood bag and their migratory potential suggest their use for temporary production of recombinant biologically active molecules to correct certain pathological conditions.This paper presents a proof-of-concept of simple,safe and effective approach for personalized ex vivo gene therapy based on gene-modified leucoconcentrate autoinfusion.The animal protocols were approved by the Kazan State Medical University Animal Care and Use Committee(approval No.5)on May 27,2014.
