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BacMam

March 16, 2024

Baculovirus gene transfer into Mammalian cells, known from scientific research articles as BacMam, is the use of baculovirus to deliver genes to mammalian cells.[1][2] Baculoviruses are insect viruses that can be modified to express proteins in mammalian cells. The unmodified baculovirus is able to enter those cells; however, its genes are not expressed unless a mammalian recognizable promoter is incorporated upstream of a gene of interest. Both unmodified baculovirus and its modified counterpart are unable to replicate in humans and are thus non-infectious.

A human mesenchymal stem cell expressing microtubule associated protein fusion to Green fluorescent protein (green) and histone 2b fusion to tagRFP (red) via BacMam gene delivery technology.

Baculovirus-mediated gene transfer was invented by Dr. Frederick M. Boyce.[3] It has gained widespread use because of advantages when compared to other transfection methods.[4][5][6] In addition, BacMam has been found to have inherent flexibility over stable cell lines,[7] which has contributed to its adoption as a standard gene transfer technique.

General properties edit

The BacMam gene delivery technology is a transient expression system, which facilitates the expression of toxic gene products. It has a broad range of transduction including many primary cell types and stem cells.[8] The baculoviral genome has a large capacity for foreign gene insertion with up to 38 kb have been tried successfully.[9] Simultaneous delivery of multiple genes to the same cell is feasible.[10] There are little to no microscopically observable cytopathic effects of BacMam particles on mammalian cells.[11] The level of gene expression can be adjusted by viral dose or chemical additions using histone deacetylase inhibitors.[12] Transduction of cells is performed by liquid-only addition and therefore BacMam is amenable to automated methods. Viruses are stable when stored at 4°C in the dark for long periods of time.[13]

Biosafety considerations edit

Baculoviruses are Risk Group 1 agents that have been widely used for over 25 years for insect cell protein production applications.[14] Baculoviruses are produced in insect cells and incapable of replicating in mammalian cells and are not known to cause disease in healthy human adults. Furthermore, BacMam viruses are inactivated by human complement, which reduces risk to researchers. Lastly, viruses used in the laboratory cannot replicate in insects so there is no environmental threat from these particles accidentally being released into the environment.[15][16]

Viral entry edit

Studies on baculovirus entry into human hepatocellular carcinoma cells suggest that BacMam enters mammalian cells via clathrin-mediated endocytosis and possibly via micropinocytosis.[17] Further studies have suggested that caveolae are somehow involved in baculovirus entry in mammalian cells.[18]

Host cell response edit

To be effective, a gene delivery technology must not interfere with normal cellular function. Cytotoxicity assays and transcriptome analyses on a human HEK cell line (HEK293) have revealed that baculovirus transduction is not cytotoxic and does not induce differential transcriptional responses.[19] Similarly, infected Schwann cells retain their characteristic morphological and molecular phenotype and are capable of differentiating in vitro and expressing the P0 myelination marker. Using complementary DNA (cDNA) microarray technology to examine in vitro and in vivo global cellular gene expression profiles in the rat brain, cultured human astrocytes , and human neuronal cells after viral transduction, host antiviral responses were observed.[20] The related genes were mainly those associated with innate immunity, including several of the genes involved in Toll-like receptor signaling pathway and cytokine-cytokine receptor interaction.

  • Bioproduction
BacMam has been used to produce proteins in large quantities using HEK293 cells in a hollow fiber bioreactor system[21]
  • High Throughput Screening
Pharmacology of G protein-coupled receptor is enabled with the use of BacMam technology in drug discovery applications[22]
  • Fluorescence Microscopy
Organelle labeling reagents are commercially available BacMam particles for labeling organelles and other subcellular structures[23]
Single mitochondrion labeled with a mitochondrial targeted green fluorescent protein[24]
  • Receptor Activation/Pathway Analysis
Characterization of serotonin receptor activation via a BacMam delivered GFP fusion to a kinase substrate[25]
  • Structural Biology
BacMam system has been used to produce soluble and membrane glycoproteins for structural studies[26]

See also edit

References edit

  1. ^ Hofmann, C; Strauss, M (1998). "Baculovirus-mediated gene transfer in the presence of human serum or blood facilitated by inhibition of the complement system". Gene Therapy. 5 (4): 531–536. doi:10.1038/sj.gt.3300607. PMID 9614578.
  2. ^ Boyce, FM; Bucher, NL (1996). "Baculovirus-mediated gene transfer into mammalian cells". Proceedings of the National Academy of Sciences of the United States of America. 93 (6): 2348–52. Bibcode:1996PNAS...93.2348B. doi:10.1073/pnas.93.6.2348. PMC 39799. PMID 8637876.
  3. ^ US patent #5,871,986
  4. ^ Kost, T; Condreay, JP (2002). "Recombinant baculoviruses as mammalian cell gene-delivery vectors". Trends in Biotechnology. 20 (4): 173–180. doi:10.1016/S0167-7799(01)01911-4. PMID 11906750.
  5. ^ Kost, Thomas A; Condreay, J Patrick; Jarvis, Donald L (2005). "Baculovirus as versatile vectors for protein expression in insect and mammalian cells". Nature Biotechnology. 23 (5): 567–575. doi:10.1038/nbt1095. PMC 3610534. PMID 15877075.
  6. ^ Kost, T; Condreay, J; Ames, R; Rees, S; Romanos, M (2007). "Implementation of BacMam virus gene delivery technology in a drug discovery setting". Drug Discovery Today. 12 (9–10): 396–403. doi:10.1016/j.drudis.2007.02.017. PMID 17467576.
  7. ^ Davenport, Elizabeth A.; Nuthulaganti, Parvathi; Ames, Robert S. (2009). "BacMam: Versatile Gene Delivery Technology for GPCR Assays". G Protein-Coupled Receptors in Drug Discovery. Methods in Molecular Biology. Vol. 552. pp. 199–211. doi:10.1007/978-1-60327-317-6_14. ISBN 978-1-60327-316-9. PMID 19513651. S2CID 21324231.
  8. ^ Zeng, Jieming; Du, Juan; Zhao, Ying; Palanisamy, Nallasivam; Wang, Shu (2007). "Baculoviral Vector-Mediated Transient and Stable Transgene Expression in Human Embryonic Stem Cells". Stem Cells. 25 (4): 1055–1061. doi:10.1634/stemcells.2006-0616. PMID 17420229.
  9. ^ Cheshenko, N; Krougliak, N; Eisensmith, R C; Krougliak, V A (2001). "A novel system for the production of fully deleted adenovirus vectors that does not require helper adenovirus". Gene Therapy. 8 (11): 846–854. doi:10.1038/sj.gt.3301459. PMID 11423932.
  10. ^ Ames, Robert; Nuthulaganti, Parvathi; Fornwald, Jim; Shabon, Usman; Van-Der-Keyl, Harjeet; Elshourbagy, Nabil (2004). "Heterologous Expression of G Protein?Coupled Receptors in U-2 OS Osteosarcoma Cells". Receptors and Channels. 10 (3–4): 117–124. doi:10.1080/10606820490515012. PMID 15512846.
  11. ^ Cheng, T; Xu, CY; Wang, YB; Chen, M; Wu, T; Zhang, J; Xia, NS (2004). "A rapid and efficient method to express target genes in mammalian cells by baculovirus". World Journal of Gastroenterology. 10 (11): 1612–8. doi:10.3748/wjg.v10.i11.1612. PMC 4572764. PMID 15162535.
  12. ^ Pfohl, JL; Worley Jf, 3rd; Condreay, JP; An, G; Apolito, CJ; Kost, TA; Truax, JF (2002). "Titration of KATP channel expression in mammalian cells utilizing recombinant baculovirus transduction". Receptors & Channels. 8 (2): 99–111. doi:10.1080/10606820212396. PMID 12448791.{{cite journal}}: CS1 maint: numeric names: authors list (link)
  13. ^ Jorio, H.; Tran, R.; Kamen, A. (2006). "Stability of Serum-Free and Purified Baculovirus Stocks under Various Storage Conditions". Biotechnology Progress. 22 (1): 319–325. doi:10.1021/bp050218v. PMID 16454526. S2CID 41069508.
  14. ^ Smith, GE; Summers, MD; Fraser, MJ (1983). "Production of human beta interferon in insect cells infected with a baculovirus expression vector". Molecular and Cellular Biology. 3 (12): 2156–65. doi:10.1128/MCB.3.12.2156. PMC 370086. PMID 6318086.
  15. ^ Kost, T. A., and Condreay, J. P. 2002a. Applied Biosafety 7:167–169.
  16. ^ Kost, T. A., Condreay, J. P., and Mickelson, C. A. 2000. Bisafety and viral gene transfer vectors. In Biological safety, principles and practices, 3rd ed. D. O. Fleming and D. L. Hunt (eds.), American Society for Microbiology Press, Washington D.C., pp. 579–597.
  17. ^ Matilainen, H.; Rinne, J.; Gilbert, L.; Marjomaki, V.; Reunanen, H.; Oker-Blom, C. (2005). "Baculovirus Entry into Human Hepatoma Cells". Journal of Virology. 79 (24): 15452–15459. doi:10.1128/JVI.79.24.15452-15459.2005. PMC 1316037. PMID 16306616.
  18. ^ Long, G.; Pan, X.; Kormelink, R.; Vlak, J. M. (2006). "Functional Entry of Baculovirus into Insect and Mammalian Cells is Dependent on Clathrin-Mediated Endocytosis". Journal of Virology. 80 (17): 8830–8833. doi:10.1128/JVI.00880-06. PMC 1563848. PMID 16912330.
  19. ^ Kenoutis, C.; Efrose, R. C.; Swevers, L.; Lavdas, A. A.; Gaitanou, M.; Matsas, R.; Iatrou, K. (2006). "Baculovirus-Mediated Gene Delivery into Mammalian Cells Does Not Alter Their Transcriptional and Differentiating Potential but is Accompanied by Early Viral Gene Expression". Journal of Virology. 80 (8): 4135–4146. doi:10.1128/JVI.80.8.4135-4146.2006. PMC 1440473. PMID 16571829.
  20. ^ Boulaire, Jérôme; Zhao, Ying; Wang, Shu (2009). "Gene expression profiling to define host response to baculoviral transduction in the brain". Journal of Neurochemistry. 109 (5): 1203–1214. doi:10.1111/j.1471-4159.2009.06015.x. PMID 19476540.
  21. ^ Jardin, B; Zhao, Y; Selvaraj, M; Montes, J; Tran, R; Prakash, S; Elias, C (2008). "Expression of SEAP (secreted alkaline phosphatase) by baculovirus mediated transduction of HEK 293 cells in a hollow fiber bioreactor system". Journal of Biotechnology. 135 (3): 272–280. doi:10.1016/j.jbiotec.2008.04.006. PMID 18499293.
  22. ^ Ames, Robert; Fornwald, James; Nuthulaganti, Parvathi; Trill, John; Foley, James; Buckley, Peter; Kost, Thomas; Wu, Zining; Romanos, Michael (2004). "BacMam Recombinant Baculoviruses in G Protein?Coupled Receptor Drug Discovery". Receptors and Channels. 10 (3–4): 99–107. doi:10.1080/10606820490514969. PMID 15512844.
  23. ^ Ames, Robert S; Kost, Thomas A; Condreay, J Patrick (2007). "BacMam technology and its application to drug discovery". Expert Opin Drug Discov. 2 (12): 1669–1681. doi:10.1517/17460441.2.12.1669. PMID 23488908. S2CID 45039136.
  24. ^ Distelmaier, Felix; Koopman, Werner J. H.; Testa, Epifania R.; De Jong, Arjan S.; Swarts, Herman G.; Mayatepek, Ertan; Smeitink, Jan A. M.; Willems, Peter H. G. M. (2008). "Life cell quantification of mitochondrial membrane potential at the single organelle level". Cytometry Part A. 73A (2): 129–138. doi:10.1002/cyto.a.20503. PMID 18163486. S2CID 5147755.
  25. ^ Huwiler, Kristin G.; MacHleidt, Thomas; Chase, Lucas; Hanson, Bonnie; Robers, Matthew B. (2009). "Characterization of serotonin 5-hydroxytryptamine-1A receptor activation using a phospho-extracellular-signal regulated kinase 2 sensor". Analytical Biochemistry. 393 (1): 95–104. doi:10.1016/j.ab.2009.06.018. PMID 19539597.
  26. ^ Dukkipati, Abhiram; Park, Hyun Ho; Waghray, Deepak; Fischer, Suzanne; Garcia, Keenan C. (2008). "BacMam system for high-level expression of recombinant soluble and membrane glycoproteins for structural studies". Protein Expr Purif. 62 (2): 160–170. doi:10.1016/j.pep.2008.08.004. PMC 2637115. PMID 18782620.

External links edit

  • The BacMam System

March 16, 2024
bacmam, this, article, contains, content, that, written, like, advertisement, please, help, improve, removing, promotional, content, inappropriate, external, links, adding, encyclopedic, content, written, from, neutral, point, view, january, 2012, learn, when,. This article contains content that is written like an advertisement Please help improve it by removing promotional content and inappropriate external links and by adding encyclopedic content written from a neutral point of view January 2012 Learn how and when to remove this template message Baculovirus gene transfer into Mammalian cells known from scientific research articles as BacMam is the use of baculovirus to deliver genes to mammalian cells 1 2 Baculoviruses are insect viruses that can be modified to express proteins in mammalian cells The unmodified baculovirus is able to enter those cells however its genes are not expressed unless a mammalian recognizable promoter is incorporated upstream of a gene of interest Both unmodified baculovirus and its modified counterpart are unable to replicate in humans and are thus non infectious A human mesenchymal stem cell expressing microtubule associated protein fusion to Green fluorescent protein green and histone 2b fusion to tagRFP red via BacMam gene delivery technology Baculovirus mediated gene transfer was invented by Dr Frederick M Boyce 3 It has gained widespread use because of advantages when compared to other transfection methods 4 5 6 In addition BacMam has been found to have inherent flexibility over stable cell lines 7 which has contributed to its adoption as a standard gene transfer technique Contents 1 General properties 2 Biosafety considerations 3 Viral entry 4 Host cell response 5 See also 6 References 7 External linksGeneral properties editThe BacMam gene delivery technology is a transient expression system which facilitates the expression of toxic gene products It has a broad range of transduction including many primary cell types and stem cells 8 The baculoviral genome has a large capacity for foreign gene insertion with up to 38 kb have been tried successfully 9 Simultaneous delivery of multiple genes to the same cell is feasible 10 There are little to no microscopically observable cytopathic effects of BacMam particles on mammalian cells 11 The level of gene expression can be adjusted by viral dose or chemical additions using histone deacetylase inhibitors 12 Transduction of cells is performed by liquid only addition and therefore BacMam is amenable to automated methods Viruses are stable when stored at 4 C in the dark for long periods of time 13 Biosafety considerations editBaculoviruses are Risk Group 1 agents that have been widely used for over 25 years for insect cell protein production applications 14 Baculoviruses are produced in insect cells and incapable of replicating in mammalian cells and are not known to cause disease in healthy human adults Furthermore BacMam viruses are inactivated by human complement which reduces risk to researchers Lastly viruses used in the laboratory cannot replicate in insects so there is no environmental threat from these particles accidentally being released into the environment 15 16 Viral entry editStudies on baculovirus entry into human hepatocellular carcinoma cells suggest that BacMam enters mammalian cells via clathrin mediated endocytosis and possibly via micropinocytosis 17 Further studies have suggested that caveolae are somehow involved in baculovirus entry in mammalian cells 18 Host cell response editTo be effective a gene delivery technology must not interfere with normal cellular function Cytotoxicity assays and transcriptome analyses on a human HEK cell line HEK293 have revealed that baculovirus transduction is not cytotoxic and does not induce differential transcriptional responses 19 Similarly infected Schwann cells retain their characteristic morphological and molecular phenotype and are capable of differentiating in vitro and expressing the P0 myelination marker Using complementary DNA cDNA microarray technology to examine in vitro and in vivo global cellular gene expression profiles in the rat brain cultured human astrocytes and human neuronal cells after viral transduction host antiviral responses were observed 20 The related genes were mainly those associated with innate immunity including several of the genes involved in Toll like receptor signaling pathway and cytokine cytokine receptor interaction BioproductionBacMam has been used to produce proteins in large quantities using HEK293 cells in a hollow fiber bioreactor system 21 High Throughput ScreeningPharmacology of G protein coupled receptor is enabled with the use of BacMam technology in drug discovery applications 22 Fluorescence MicroscopyOrganelle labeling reagents are commercially available BacMam particles for labeling organelles and other subcellular structures 23 Single mitochondrion labeled with a mitochondrial targeted green fluorescent protein 24 Receptor Activation Pathway AnalysisCharacterization of serotonin receptor activation via a BacMam delivered GFP fusion to a kinase substrate 25 Structural BiologyBacMam system has been used to produce soluble and membrane glycoproteins for structural studies 26 See also editBaculovirusReferences edit Hofmann C Strauss M 1998 Baculovirus mediated gene transfer in the presence of human serum or blood facilitated by inhibition of the complement system Gene Therapy 5 4 531 536 doi 10 1038 sj gt 3300607 PMID 9614578 Boyce FM Bucher NL 1996 Baculovirus mediated gene transfer into mammalian cells Proceedings of the National Academy of Sciences of the United States of America 93 6 2348 52 Bibcode 1996PNAS 93 2348B doi 10 1073 pnas 93 6 2348 PMC 39799 PMID 8637876 US patent 5 871 986 Kost T Condreay JP 2002 Recombinant baculoviruses as mammalian cell gene delivery vectors Trends in Biotechnology 20 4 173 180 doi 10 1016 S0167 7799 01 01911 4 PMID 11906750 Kost Thomas A Condreay J Patrick Jarvis Donald L 2005 Baculovirus as versatile vectors for protein expression in insect and mammalian cells Nature Biotechnology 23 5 567 575 doi 10 1038 nbt1095 PMC 3610534 PMID 15877075 Kost T Condreay J Ames R Rees S Romanos M 2007 Implementation of BacMam virus gene delivery technology in a drug discovery setting Drug Discovery Today 12 9 10 396 403 doi 10 1016 j drudis 2007 02 017 PMID 17467576 Davenport Elizabeth A Nuthulaganti Parvathi Ames Robert S 2009 BacMam Versatile Gene Delivery Technology for GPCR Assays G Protein Coupled Receptors in Drug Discovery Methods in Molecular Biology Vol 552 pp 199 211 doi 10 1007 978 1 60327 317 6 14 ISBN 978 1 60327 316 9 PMID 19513651 S2CID 21324231 Zeng Jieming Du Juan Zhao Ying Palanisamy Nallasivam Wang Shu 2007 Baculoviral Vector Mediated Transient and Stable Transgene Expression in Human Embryonic Stem Cells Stem Cells 25 4 1055 1061 doi 10 1634 stemcells 2006 0616 PMID 17420229 Cheshenko N Krougliak N Eisensmith R C Krougliak V A 2001 A novel system for the production of fully deleted adenovirus vectors that does not require helper adenovirus Gene Therapy 8 11 846 854 doi 10 1038 sj gt 3301459 PMID 11423932 Ames Robert Nuthulaganti Parvathi Fornwald Jim Shabon Usman Van Der Keyl Harjeet Elshourbagy Nabil 2004 Heterologous Expression of G Protein Coupled Receptors in U 2 OS Osteosarcoma Cells Receptors and Channels 10 3 4 117 124 doi 10 1080 10606820490515012 PMID 15512846 Cheng T Xu CY Wang YB Chen M Wu T Zhang J Xia NS 2004 A rapid and efficient method to express target genes in mammalian cells by baculovirus World Journal of Gastroenterology 10 11 1612 8 doi 10 3748 wjg v10 i11 1612 PMC 4572764 PMID 15162535 Pfohl JL Worley Jf 3rd Condreay JP An G Apolito CJ Kost TA Truax JF 2002 Titration of KATP channel expression in mammalian cells utilizing recombinant baculovirus transduction Receptors amp Channels 8 2 99 111 doi 10 1080 10606820212396 PMID 12448791 a href Template Cite journal html title Template Cite journal cite journal a CS1 maint numeric names authors list link Jorio H Tran R Kamen A 2006 Stability of Serum Free and Purified Baculovirus Stocks under Various Storage Conditions Biotechnology Progress 22 1 319 325 doi 10 1021 bp050218v PMID 16454526 S2CID 41069508 Smith GE Summers MD Fraser MJ 1983 Production of human beta interferon in insect cells infected with a baculovirus expression vector Molecular and Cellular Biology 3 12 2156 65 doi 10 1128 MCB 3 12 2156 PMC 370086 PMID 6318086 Kost T A and Condreay J P 2002a Applied Biosafety 7 167 169 Kost T A Condreay J P and Mickelson C A 2000 Bisafety and viral gene transfer vectors In Biological safety principles and practices 3rd ed D O Fleming and D L Hunt eds American Society for Microbiology Press Washington D C pp 579 597 Matilainen H Rinne J Gilbert L Marjomaki V Reunanen H Oker Blom C 2005 Baculovirus Entry into Human Hepatoma Cells Journal of Virology 79 24 15452 15459 doi 10 1128 JVI 79 24 15452 15459 2005 PMC 1316037 PMID 16306616 Long G Pan X Kormelink R Vlak J M 2006 Functional Entry of Baculovirus into Insect and Mammalian Cells is Dependent on Clathrin Mediated Endocytosis Journal of Virology 80 17 8830 8833 doi 10 1128 JVI 00880 06 PMC 1563848 PMID 16912330 Kenoutis C Efrose R C Swevers L Lavdas A A Gaitanou M Matsas R Iatrou K 2006 Baculovirus Mediated Gene Delivery into Mammalian Cells Does Not Alter Their Transcriptional and Differentiating Potential but is Accompanied by Early Viral Gene Expression Journal of Virology 80 8 4135 4146 doi 10 1128 JVI 80 8 4135 4146 2006 PMC 1440473 PMID 16571829 Boulaire Jerome Zhao Ying Wang Shu 2009 Gene expression profiling to define host response to baculoviral transduction in the brain Journal of Neurochemistry 109 5 1203 1214 doi 10 1111 j 1471 4159 2009 06015 x PMID 19476540 Jardin B Zhao Y Selvaraj M Montes J Tran R Prakash S Elias C 2008 Expression of SEAP secreted alkaline phosphatase by baculovirus mediated transduction of HEK 293 cells in a hollow fiber bioreactor system Journal of Biotechnology 135 3 272 280 doi 10 1016 j jbiotec 2008 04 006 PMID 18499293 Ames Robert Fornwald James Nuthulaganti Parvathi Trill John Foley James Buckley Peter Kost Thomas Wu Zining Romanos Michael 2004 BacMam Recombinant Baculoviruses in G Protein Coupled Receptor Drug Discovery Receptors and Channels 10 3 4 99 107 doi 10 1080 10606820490514969 PMID 15512844 Ames Robert S Kost Thomas A Condreay J Patrick 2007 BacMam technology and its application to drug discovery Expert Opin Drug Discov 2 12 1669 1681 doi 10 1517 17460441 2 12 1669 PMID 23488908 S2CID 45039136 Distelmaier Felix Koopman Werner J H Testa Epifania R De Jong Arjan S Swarts Herman G Mayatepek Ertan Smeitink Jan A M Willems Peter H G M 2008 Life cell quantification of mitochondrial membrane potential at the single organelle level Cytometry Part A 73A 2 129 138 doi 10 1002 cyto a 20503 PMID 18163486 S2CID 5147755 Huwiler Kristin G MacHleidt Thomas Chase Lucas Hanson Bonnie Robers Matthew B 2009 Characterization of serotonin 5 hydroxytryptamine 1A receptor activation using a phospho extracellular signal regulated kinase 2 sensor Analytical Biochemistry 393 1 95 104 doi 10 1016 j ab 2009 06 018 PMID 19539597 Dukkipati Abhiram Park Hyun Ho Waghray Deepak Fischer Suzanne Garcia Keenan C 2008 BacMam system for high level expression of recombinant soluble and membrane glycoproteins for structural studies Protein Expr Purif 62 2 160 170 doi 10 1016 j pep 2008 08 004 PMC 2637115 PMID 18782620 External links editThe BacMam System Retrieved from https en wikipedia org w index php title BacMam amp oldid 1213163546, wikipedia, wiki, book, books, library,

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