TK (His) Lentiviral particles
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1. 7930 Arjons Drive Suite B San Diego CA 92126 el arue ne Phone 858 6788683 Fax 800 3804198 Email orders gentarget com Pre made Lentiviral Particles for Thymidine kinase manual Product Name TK His Lentiviral particles Cat LVPO18 Amount 200ul 1 x 10 IFU mL in DMEM containing 10 FBS and 60ug ml ploybrene 10x Storage lt 70 C avoid repeat freeze thaw cycles Product Description Lentiviral particles or lentivirus is a gene delivery tool produced from lentivectors for gene expression or knockdown GenTarget s lentivector system is Human Immunodeficiency Virus 1 HIV based plasmids for gene expression and knockdown The lentivectors are used to generate lentiviral particles lentivirus that can be transduced into almost all kinds of mammalian cells including stem cells primary cells and non dividing cells both in vivo and in vitro Lentiviral Particles stably integrate into the transduced cells genome for long term expression making it a great gene transfer agent Pre made Thymidine kinase TK lentiviral particles are generated from GenTarget s re engineered lentivector system TK gene was fully sequence verified expressed under suCMV promoter with N terminal 6His tag VSV G pseudotyped lentiviral particles are generated in 293T cell and packed in 10 of FBS in DMEM supplied as 200ul per vial at 1 x 10 IFU mI containing 60ug ml 10x of polybrene See FAQ for pre made lentiviral particle2. ells Grow cells in complete suspension culture medium use a shaking flask in a CO incubator if necessary Measure cell density When density has reached 3 x 10 cells ml measured viability should be gt 90 Dilute cells into 1 x 10 cell ml in complete medium Day 1 e Thaw lentiviral particles at room temperature e Add premade lentiviral particles into the diluted cells at a ratio of 50 to 100 ul virus per 0 5 ml of cells Note depending on cell type you may need to use more or less virus e Grow cells in a shaking flask in a CO2 incubator Day 2 At 24 hours after transduction add an equal amount of fresh medium containing relevant antibiotics Note amount of antibiotic depends on cell type Continue growing cells in CO2 incubator Day 3 At 72 hours after transduction check fluorescence with a fluorescence microscope or calculate the transduction efficiency using a cell sorter such as FACS or Guava Sort for fluorescence positive cells and maintain antibiotic selection to generate a stable cell line Note Filter wavelength settings GFP filter Ex450 490 Em525 RFP filter Ex545 Em620 Safety Precaution Gentarget lentiviral particles adapts must advanced lentiviral safety features using the third generation vectors with self inactivation SIN 3UTR and the premade lentivirus is replication incompetent However please use extra caution when using lentiviral particles Use the lentiviral particles in Bio safe
3. et Inc Copyrights 2015 7930 Arjons Drive Suite B San Diego CA 92126 el arge ne Phone 858 6788683 Fax 800 3804198 Email orders gentarget com lentivirus microRNA Premade lentivirus expression human or mouse precursor and anti miRNA And anti miRNA lentivector and virus for human and microRNA mouse miRNA lentivirus Negative Premade negative control lentivirus with different markers control serves as the negative control of lentivurs treatment for lentiviruses validation of the specificity of any lentivirus target expression effects Other Ready to use lentivirus expressing specific enzymes with Enzyme different selection markers expression References ar NIH stem cell training program Link 2 Takahashi K and Yamanaka S 2006 Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors Cell 126 663 676 3 Yu J Vodyanik M A Smuga Otto K Antosiewicz Bourget J Frane J L Tian S Nie J Jonsdottir G A Ruotti V Stewart R Slukvin l l and Thomson J A 2007 Induced pluripotent stem cell lines derived from human somatic cells Science 318 1917 1920 4 Park I H et al Reprogramming of human somatic cells to pluripotency with defined factors Nature 2008 451 7175 p 141 6 5 Shao L et al Generation of iPS cells using defined factors linked via the self cleaving 2A sequences in a single open
4. followed by antibiotic selection Day O Seed cells in complete medium at the appropriate density and incubate overnight Note at the time of transduction cells should be 50 75 confluent For example seed HeLa cells at 0 5 x 10 ml x 0 5m in a well of a 24 well plate Day 1 e Remove the culture medium and add 0 5ml fresh warm complete medium e Thaw the pre made lentiviral stock at room temperature and add the appropriate amount of virus stock to obtain the desired MOI e Return cells to 37 C CO2 incubator Note Try to avoid freezing and thawing If you do not use all of the virus at one time you may re freeze the virus at 80 C for future use virus titer will decrease by 10 for each freeze thaw cycle Day 3 At 72hr after transduction check the transduction rate by fluorescence microscopy or calculate the exact transduction rate by flow cytometry FACS or Guava Day 3 optional Sort transduced cells by FACS and select for antibiotic resistance A pilot experiment should be done to determine the antibiotic s kill curve Pre made Lentiviral Particles for TK Product manual Page 2 of 5 www gentarget com GenTarget Inc Copyrights 2015 7930 Arjons Drive Suite B San Diego CA 92126 el arue ne Phone 858 6788683 Fax 800 3804198 Email orders gentarget com for your specific cell line refer to the pertinent literature on generation of stable cell lines 2 Transduction Protocol for Suspension C
5. reading frame Cell Res 2009 19 3 p 296 306 6 NIH Guidelines for Biosafety Considerations for Research with Lentiviral Vectors Link 7 CDC guidelines for Lab Biosafety levels Link Pre made Lentiviral Particles for TK Product manual Page 5 of 5 www gentarget com GenTarget Inc Copyrights 2015
6. s pdf for details Key features 1 Lentiviral particles contain RFP blasticidin resistant gene used for generating stable cell lines by Blasticidin antibiotics or via fluorescent cell sorting 2 Target was expressed with a N term His tag for purification of target protein if desired 3 The strongest suCMV promoter make the pre made virus a ideal tool for mammalian protein expression stable cell line construction and enzymatic assays both in vivo or in vitro see schematic vector map below 4 The lentivirus are ready and easy to use simply add 50ul into your cell culture Pre made Lentiviral Particles for TK Product manual Page 1 of 5 www gentarget com GenTarget Inc Copyrights 2015 7930 Arjons Drive Suite B San Diego CA 92126 el arue ne Phone 858 6788683 Fax 800 3804198 Email orders gentarget com TK TK lentivector e Rsv RFP Bsd meneg SL Transduction Protocols 1 Transduction Protocol for Adhesive cells Note Pre made lentivirus is provided ready to use so it can be simply added into your cell culture the amount of virus to add depends on cell type For quick transduction add 50 ul of virus into each well of 24 well plate where cell density is 50 to 75 After 72 hours no need to change medium visualize positive transduction rate by fluorescence microscopy For stable cell line generation pass cells into medium containing antibiotic or perform fluorescence cell sorting
7. selection markers CRE Premade lentivirus for expressing nuclear permeant CRE recombinase recombinase with different flurescent and antibiotic markers LoxP Premade lentivirus expressing LoxP GFP Stop LoxP RFP ColorSwitch cassette used to monitor the CRE recombination event in vivo CRISPR hu Preamde lentivirus express humanzied wild type Cas9 CAS9 endonuclease for genomic editing with CRISPR TetR Premade lentivirus expressin TetR tetracycline regulator protein inducible the repressor protein for the inducible expression system expression repressor Premde lentivirus for human and mouse iPS Myc NANOG iPS factors OCT4 SOX2 FLF4 factors with different fluorescent and antibitoic markers T antigen Express SV40 large T antigen with different selection markers Expression Cell Premade lentivirus for cell organelle imaging The fluorescent Organelle marker GFP RFP CFP was sub cellular localized in different imaging cell organelle for living cell imaging LacZ Express different full length B galactosidase lacZ with different expression selection markers Anti miNA Pre made lentivirus expression a specific anti miRNA cassette lentivirus Fluorescent Pre made lentivirus expression a GFP RFP CFP ORF fusion ORF fusion target Pre made Premade shRNA lentivirus for knockdown a specific genes P53 shRNA LacZ Luciferase and more Pre made Lentiviral Particles for TK Product manual Page 4 of 5 www gentarget com GenTarg
8. ty II cabinet Wear glove all the time when handling Lentiviral particles Please refer CDC and NIH s guidelines for more details regarding to safety issues Warranty This product is for research use only It is warranted to meet its quality as described when used in accordance with its instructions GenTarget disclaims any implied warranty of this product for particular application In no event shall Pre made Lentiviral Particles for TK Product manual Page 3 of 5 www gentarget com GenTarget Inc Copyrights 2015 GenTarget be liable for any incidental or consequential damages in connection with the products GenTarget s sole remedy for breach of this warranty should be at GenTarset inc 7930 Arjons Drive Suite B San Diego CA 92126 Phone 858 6788683 Fax 800 3804198 Email orders gentarget com GenTarget s option to replace the products Related Products GenTarget s pre made lentivirus product category Product Product Description Category please click category name to see product s pages Human Premade lentivirus expressin a human mouse or rat gene with mouse or rat RFP Blastididin fusion dual markers ORFs Fluorescent Preamde lentivirus express human codon optimized fluorescent markers protein GFP RFP CFP BFP YFP Luciferase Premade lentivirus for all kinds of luciferase protein expression expression firefly and Renilla with different antibiotic
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