Lentiviral vectors with fluorescent proteins

The pLVX-AcGFP1-C1 vector is an HIV-1 based lentiviral expression vector that can produce recombinant lentivirus that efficiently transduces both dividing and non-dividing mammalian cells. This vector expresses a variant of the green fluorescent protein (GFP) from Aequorea coerulescens. pLVX-AcGFP1-C1 allows cloning of a cDNA into the multiple cloning site (MCS) downstream of the coding sequence for AcGFP1. The cDNA is subsequently expressed as a fusion to the C-terminus of the AcGFP1 protein. Thus,
this vector allows the expression of fluorescent fusion proteins in difficult-to-transfect cells. It also contains a puromycin selection marker for selection of stable transductants. The unmodified vector will express AcGFP1 protein and may be used to produce marker virus to optimize infection protocols.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

The pLVX-AcGFP1-N1 vector is an HIV-1-based lentiviral expression vector that can produce recombinant lentivirus that efficiently transduces both dividing and non-dividing mammalian cells. This vector expresses a variant of the green fluorescent protein (GFP) from Aequorea coerulescens. pLVX-AcGFP1-N1 allows cloning of a cDNA into the multiple cloning site (MCS) upstream of the coding sequence for AcGFP1. The cDNA is subsequently expressed as a fusion to the N-terminus of the AcGFP1 protein. Thus, this vector allows the expression of fluorescent fusion proteins in difficult-to-transfect cells. It also contains a puromycin selection marker for selection of stable transductants. The unmodified vector will express AcGFP1 protein and may be used to produce marker virus to optimize infection protocols.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

This lentiviral expression vector encodes an AmCyan1 fluorescent protein tag. The protein is a human codon-optimized variant of the Anemonia majano cyan fluorescent protein engineered for increased brightness and higher expression in mammalian cells. Inserting a cDNA in the MCS downstream of the AmCyan1 coding sequence joins your protein of interest to the C-terminus of AmCyan1, and allows it to be tracked and studied in transduced cells. To package the vector into high-titer, replication-incompetent lentivirus, we recommend using the Lenti-X HTX Packaging System (Cat. No. 631247) and the Lenti-X 293T Cell Line (Cat. No. 632180). The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

This lentiviral expression vector encodes an AmCyan1 fluorescent protein tag. The protein is a human codon-optimized variant of the Anemonia majano cyan fluorescent protein engineered for increased brightness and higher expression in mammalian cells. Inserting a cDNA in the MCS upstream of the AmCyan1 coding sequence joins your protein of interest to the N-terminus of AmCyan1, and allows it to be tracked and studied in transduced cells. To package the vector into high-titer, replication-incompetent lentivirus, we recommend using the Lenti-X HTX Packaging System (Cat. No. 631247) and the Lenti-X 293T Cell Line (Cat. No. 632180). The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

This lentiviral expression vector encodes a DsRed-Express2 fluorescent protein tag. The protein is a rapidly maturing tetrameric fluorescent protein from Discosoma sp. that has minimal cytotoxicity, high solubility, and exhibits strong and stable expression in mammalian cells. Inserting a cDNA in the MCS downstream of the DsRed-Express2 coding sequence joins your protein of interest to the C-terminus of the tag, and allows the fusion protein to be tracked and studied in transduced cells. To package the vector into high-titer, replication-incompetent lentivirus, we recommend using the Lenti-X Packaging Single Shots (VSV-G) and the Lenti-X 293T Cell Line. The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

This lentiviral expression vector encodes a DsRed-Express2 fluorescent protein tag. The protein is a rapidly maturing tetrameric fluorescent protein from Discosoma sp. that has minimal cytotoxicity, high solubility, and exhibits strong and stable expression mammalian cells. Inserting a cDNA in the MCS upstream of the DsRed-Express2 coding sequence joins your protein of interest to the N-terminus of the tag, and allows the fusion protein to be tracked and studied in transduced cells. To package the vector into high-titer, replication-incompetent lentivirus, we recommend using the Lenti-X Packaging Single Shots and the Lenti-X 293T Cell Line. The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

The pLVX-DsRed-Monomer-C1 vector is an HIV-1-based lentiviral expression vector that can produce lentivirus that efficiently transduces both dividing and non-dividing mammalian cells. This vector expresses a monomeric mutant of the Discosoma sp. red fluorescent protein (DsRed). pLVX-DsRed-Monomer-C1 allows cloning of a cDNA into the multiple cloning site (MCS) downstream of the DsRed-Monomer coding sequence. The cDNA is subsequently expressed as a fusion to the C-terminus of the DsRed-Monomer protein. Thus, this vector allows the expression of fluorescent fusion proteins in difficult-to-transfect cells. It also contains a puromycin selection marker for selection of stable transductants. The unmodified vector will express DsRed-Monomer protein and may be used to produce marker virus to optimize infection protocols.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

The pLVX-DsRed-Monomer-N1 vector is an HIV-1-based lentiviral expression vector that can produce recombinant lentivirus that efficiently transduces both dividing and non-dividing mammalian cells. This vector expresses a monomeric mutant of the Discosoma sp. red fluorescent protein (DsRed). pLVX-DsRed-Monomer-N1 allows cloning of a cDNA into the multiple cloning site (MCS) upstream of the DsRed-Monomer coding sequence. The cDNA is subsequently expressed as a fusion to the N-terminus of the DsRed-Monomer protein. Thus, this vector allows the expression of fluorescent fusion proteins in difficult-to-transfect cells. It also contains a puromycin selection marker for selection of stable transductants. The unmodified vector will express DsRed-Monomer protein and may be used to produce marker virus to optimize infection protocols.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

The pLVX-IRES-mCherry Vector is a bicistronic lentiviral expression vector that can be used to generate high-titer lentivirus for transducing dividing or nondividing mammalian cells. The vector contains an internal ribosomal entry site (IRES) which allows a gene-of-interest and the mCherry fluorescent protein to be simultaneously coexpressed from a single mRNA transcript. When used with Lenti-X Packaging Single Shots and the Lenti-X 293T Cell Line (Cat. No. 632180), the vector generates high titers of replication-incompetent, VSV-G-pseudotyped lentivirus.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

Required Products

Cat. #	Product	Size	Price
631232	Lenti-X™ Concentrator	500 mL	$719.00
The Lenti-X Concentrator is a complete reagent for the concentration of infectious lentiviral vector particles. This reagent provides a scalable alternative to ultracentrifugation for viral particle concentration. Vector supernatants can be concentrated 10–100 fold, depending upon the volumes used. Documents Components Image Data Back Rapid lentivirus concentration Rapid lentivirus concentration. Add Lenti-X Concentrator reagent to clarified viral supernatant, incubate for 30 min to overnight at 4°C, and spin. That’s it. Back Lentiviral concentration with high yield Lentiviral concentration with high yield. Lentiviral supernatant from a pLVX-ZsGreen1 vector was concentrated from 3 ml down to 30 µl using the Lenti-X Concentrator Reagent. The titer before and after concentration were measured and confirmed that >90% of the lentivirus remained functional. Samples were titrated on HT1080 cells and analyzed by flow cytometry 48 hr post-transduction. Back Concentrate lentivirus from any volume or from any lentiviral titer Concentrate lentivirus from any volume or from any lentiviral titer. Lentiviral supernatant was diluted into 250 ml and then concentrated down to 2.5 ml using Lenti-X Concentrator (Panel A). 10-fold serial dilutions of a high-titer lentiviral supernatant (high, medium, and low) were concentrated from a volume of 10 ml down to 100 µl using the Lenti-X Concentrator Reagent (Panel B). Titrations were performed using HT1080 cells and flow cytometry 48 hr post-transduction. Back Lenti-X Concentrator Lenti-X Concentrator. Catalog # 631232.
631275	Lenti-X™ Packaging Single Shots (VSV-G)	16 Rxns	$795.00
Lenti-X Packaging Single Shots (VSV-G) provide an extremely simple and consistent one-step method for producing high-titer lentivirus. No additional transfection reagent is needed because Lenti-X Packaging Single Shots (VSV-G) consist of pre-aliquoted, lyophilized, single tubes of Xfect Transfection Reagent premixed with an optimized formulation of VSV-G pseudotyped Lenti-X lentiviral packaging plasmids. High-titer virus is produced by simply reconstituting this mixture with your lentiviral vector of choice in sterile water and adding it to 293T cells, e.g., Lenti-X 293T Cells (Cat. No. 632180), in a 10 cm dish. Documents Components You May Also Like Image Data Back The Lenti-X Packaging Single Shots (VSV-G) protocol The Lenti-X Packaging Single Shots (VSV-G) protocol. Back Consistent, high-efficiency transfections lead to high titers Consistent, high-efficiency transfections lead to high titers. A lentiviral vector containing the ZsGreen1 gene was packaged according to the Lenti-X single shots protocol in four independent experiments. Briefly, 7 µg of pLVX-ZsGreen1 plasmid was added to each of four Lenti-X single shots; the tubes were vortexed for 20 sec and incubated at room temperature for 10 min. Then, the mixture was added to cultured Lenti-X 293T cells that were approximately 80% confluent. 48 hours after transfection, the cells were imaged by fluorescence microscopy (Panel A, top) and light microscopy (Panel A, bottom). After images were taken, the supernatant was harvested and used infect HT1080 cells for titer determination (Panel B, IFU/ml). Back High-titer virus was produced regardless of the lentiviral vector backbone with Lenti-X packaging single shots High-titer virus was produced regardless of the lentiviral vector backbone with Lenti-X packaging single shots. A CMV ZsGreen1 expression cassette was cloned into several lentiviral vector backbones. These vectors were then packaged into lentivirus using the Lenti-X packaging single shots following the provided protocol. Briefly, 7 µg of pLVX-ZsGreen1 plasmid was added to each of four Lenti-X single shots, and the tubes were vortexed for 20 sec and incubated at room temperature for 10 min. Then, the mixture was added to cultured Lenti-X 293T cells that were approximately 80% confluent. After 48 hours, titer was determined using several methods. To determine infectivity, the supernatant was harvested and used to infect HT1080 cells (Flow Cytometry). Harvested viral supernatants were also analyzed by RT-PCR to quantify viral genome copies (qRT-PCR, Lenti-X qRT-PCR Titration Kit), ELISA to measure p24 (p24 ELISA, Lenti-X p24 Rapid Titer Kit), and by a rapid lentiviral detection method (Lenti-X GoStix). Back A comparison of fourth- and third-generation lentiviral packaging systems A comparison of fourth- and third-generation lentiviral packaging systems . Our Lenti-X Packaging single shots utilize a packaging system that consists of five separate components (Panel A), mixed in proprietary proportions for optimized packaging activity. The separation of the gag, pol, and env genes effectively reduces the incidence of RCL (Wu et al., 2000). High levels of expression of essential viral components are driven by the Tet-Off and Tat transactivators, which induce a cascade of expression that results in high titers of lentivirus. The pol gene is fused to vpr to ensure transport of the reverse transcriptase/integrase protein into the recombinant lentiviral particle. Not all vector elements are shown. Other 3rd generation lentiviral packaging systems (Panel B) generate lower titers, do not contain separate gag and pol sequences, and do not use a transactivation cascade mechanism.

The pLVX-IRES-tdTomato Vector is a bicistronic lentiviral expression vector that can be used to generate high-titer lentivirus for transducing dividing or nondividing mammalian cells. The vector contains an internal ribosomal entry site (IRES) which allows a gene-of-interest and the tdTomato fluorescent protein to be simultaneously coexpressed from a single mRNA transcript. When used with the Lenti-X Packaging Single Shots and the Lenti-X 293T Cell Line (Cat. No. 632180), the vector generates high titers of replication-incompetent, VSV-G-pseudotyped lentivirus.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

Required Products

Cat. #	Product	Size	Price
632180	Lenti-X™ 293T Cell Line	1 mL	$307.00
The Lenti-X 293T Cell Line is a subclone of the transformed human embryonic kidney cell line, HEK 293, which is highly transfectable and supports high levels of viral protein expression. When transfected with the Lenti-X HTX Packaging System and a lentiviral vector, these cells are capable of producing lentiviral titers as high as >10⁸ ifu/ml, as determined by flow cytometry. The cell line also constitutively expresses the simian virus 40 (SV40) large T antigen. Documents Components You May Also Like Image Data Resources Back Lenti-X 293T Cell Line Lenti-X 293T Cell Line. Cat. # 632180. Back Clontech's lentiviral packaging system (Panel A) and a lentiviral packaging system from a leading competitor (Panel B) were each used to generate viral supernatants from their respective lentiviral system vectors that were engineered to express the ZsGreen1 fluorescent protein Clontech's lentiviral packaging system (Panel A) and a lentiviral packaging system from a leading competitor (Panel B) were each used to generate viral supernatants from their respective lentiviral system vectors that were engineered to express the ZsGreen1 fluorescent protein. As little as 10 µl of supernatant from Lenti-X transduced the majority of these HeLa cells, whereas 10 µl of supernatant from the other system transduced only a small percentage of the cells. Transduced cells were quantified by flow cytometry. Back Transduction of neural progenitor cells by Lenti-X lentivirus Transduction of neural progenitor cells by Lenti-X lentivirus. Recombinant lentivirus for expressing ZsGreen1 was produced using Lenti-X virus and used to transduce normal human neural progenitor cells. A single transduced cell is shown under phase contrast microscopy (Panel A) and fluorescence microscopy (Panel B). Back High-titer lentivirus production High-titer lentivirus production. Lenti-X 293T cells were transduced with the indicated volumes (µl) of lentiviral packaging supernatant generated with the Lenti-X Expression System and then selected with puromycin for 9 days to allow the formation of the resistant colonies, which were then stained with crystal violet. Back 293T cell line for higher titers 293T cell line for higher titers. We used our fourth-generation lentiviral packaging system and one of our pLVX-lentiviral vectors to compare the virus production of the Lenti-X 293T Cell Line to that of two other commonly used HEK 293-based cell lines. Lenti-X 293T cells clearly outperformed the other cell lines—producing over 6X more virus than 293FT cells and up to 30X more virus than the parental HEK 293 cell line.
631275	Lenti-X™ Packaging Single Shots (VSV-G)	16 Rxns	$795.00
Lenti-X Packaging Single Shots (VSV-G) provide an extremely simple and consistent one-step method for producing high-titer lentivirus. No additional transfection reagent is needed because Lenti-X Packaging Single Shots (VSV-G) consist of pre-aliquoted, lyophilized, single tubes of Xfect Transfection Reagent premixed with an optimized formulation of VSV-G pseudotyped Lenti-X lentiviral packaging plasmids. High-titer virus is produced by simply reconstituting this mixture with your lentiviral vector of choice in sterile water and adding it to 293T cells, e.g., Lenti-X 293T Cells (Cat. No. 632180), in a 10 cm dish. Documents Components You May Also Like Image Data Back The Lenti-X Packaging Single Shots (VSV-G) protocol The Lenti-X Packaging Single Shots (VSV-G) protocol. Back Consistent, high-efficiency transfections lead to high titers Consistent, high-efficiency transfections lead to high titers. A lentiviral vector containing the ZsGreen1 gene was packaged according to the Lenti-X single shots protocol in four independent experiments. Briefly, 7 µg of pLVX-ZsGreen1 plasmid was added to each of four Lenti-X single shots; the tubes were vortexed for 20 sec and incubated at room temperature for 10 min. Then, the mixture was added to cultured Lenti-X 293T cells that were approximately 80% confluent. 48 hours after transfection, the cells were imaged by fluorescence microscopy (Panel A, top) and light microscopy (Panel A, bottom). After images were taken, the supernatant was harvested and used infect HT1080 cells for titer determination (Panel B, IFU/ml). Back High-titer virus was produced regardless of the lentiviral vector backbone with Lenti-X packaging single shots High-titer virus was produced regardless of the lentiviral vector backbone with Lenti-X packaging single shots. A CMV ZsGreen1 expression cassette was cloned into several lentiviral vector backbones. These vectors were then packaged into lentivirus using the Lenti-X packaging single shots following the provided protocol. Briefly, 7 µg of pLVX-ZsGreen1 plasmid was added to each of four Lenti-X single shots, and the tubes were vortexed for 20 sec and incubated at room temperature for 10 min. Then, the mixture was added to cultured Lenti-X 293T cells that were approximately 80% confluent. After 48 hours, titer was determined using several methods. To determine infectivity, the supernatant was harvested and used to infect HT1080 cells (Flow Cytometry). Harvested viral supernatants were also analyzed by RT-PCR to quantify viral genome copies (qRT-PCR, Lenti-X qRT-PCR Titration Kit), ELISA to measure p24 (p24 ELISA, Lenti-X p24 Rapid Titer Kit), and by a rapid lentiviral detection method (Lenti-X GoStix). Back A comparison of fourth- and third-generation lentiviral packaging systems A comparison of fourth- and third-generation lentiviral packaging systems . Our Lenti-X Packaging single shots utilize a packaging system that consists of five separate components (Panel A), mixed in proprietary proportions for optimized packaging activity. The separation of the gag, pol, and env genes effectively reduces the incidence of RCL (Wu et al., 2000). High levels of expression of essential viral components are driven by the Tet-Off and Tat transactivators, which induce a cascade of expression that results in high titers of lentivirus. The pol gene is fused to vpr to ensure transport of the reverse transcriptase/integrase protein into the recombinant lentiviral particle. Not all vector elements are shown. Other 3rd generation lentiviral packaging systems (Panel B) generate lower titers, do not contain separate gag and pol sequences, and do not use a transactivation cascade mechanism.

The pLVX-IRES-ZsGreen1 Vector is a bicistronic lentiviral expression vector that can be used to generate high-titer lentivirus for transducing dividing or non-dividing mammalian cells. The vector contains an internal ribosomal entry site (IRES) which allows a gene-of-interest and the ZsGreen1 fluorescent protein to be simultaneously coexpressed from a single mRNA transcript. When used with Lenti-X Packaging Single Shots (e.g. Cat. No. 631275) and our Lenti-X 293T Cell Line (Cat. No. 632180), the vector generates high titers of replication-incompetent, VSV-G pseudotyped lentivirus.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

Required Products

Cat. #	Product	Size	Price
632180	Lenti-X™ 293T Cell Line	1 mL	$307.00
The Lenti-X 293T Cell Line is a subclone of the transformed human embryonic kidney cell line, HEK 293, which is highly transfectable and supports high levels of viral protein expression. When transfected with the Lenti-X HTX Packaging System and a lentiviral vector, these cells are capable of producing lentiviral titers as high as >10⁸ ifu/ml, as determined by flow cytometry. The cell line also constitutively expresses the simian virus 40 (SV40) large T antigen. Documents Components You May Also Like Image Data Resources Back Lenti-X 293T Cell Line Lenti-X 293T Cell Line. Cat. # 632180. Back Clontech's lentiviral packaging system (Panel A) and a lentiviral packaging system from a leading competitor (Panel B) were each used to generate viral supernatants from their respective lentiviral system vectors that were engineered to express the ZsGreen1 fluorescent protein Clontech's lentiviral packaging system (Panel A) and a lentiviral packaging system from a leading competitor (Panel B) were each used to generate viral supernatants from their respective lentiviral system vectors that were engineered to express the ZsGreen1 fluorescent protein. As little as 10 µl of supernatant from Lenti-X transduced the majority of these HeLa cells, whereas 10 µl of supernatant from the other system transduced only a small percentage of the cells. Transduced cells were quantified by flow cytometry. Back Transduction of neural progenitor cells by Lenti-X lentivirus Transduction of neural progenitor cells by Lenti-X lentivirus. Recombinant lentivirus for expressing ZsGreen1 was produced using Lenti-X virus and used to transduce normal human neural progenitor cells. A single transduced cell is shown under phase contrast microscopy (Panel A) and fluorescence microscopy (Panel B). Back High-titer lentivirus production High-titer lentivirus production. Lenti-X 293T cells were transduced with the indicated volumes (µl) of lentiviral packaging supernatant generated with the Lenti-X Expression System and then selected with puromycin for 9 days to allow the formation of the resistant colonies, which were then stained with crystal violet. Back 293T cell line for higher titers 293T cell line for higher titers. We used our fourth-generation lentiviral packaging system and one of our pLVX-lentiviral vectors to compare the virus production of the Lenti-X 293T Cell Line to that of two other commonly used HEK 293-based cell lines. Lenti-X 293T cells clearly outperformed the other cell lines—producing over 6X more virus than 293FT cells and up to 30X more virus than the parental HEK 293 cell line.
631275	Lenti-X™ Packaging Single Shots (VSV-G)	16 Rxns	$795.00
Lenti-X Packaging Single Shots (VSV-G) provide an extremely simple and consistent one-step method for producing high-titer lentivirus. No additional transfection reagent is needed because Lenti-X Packaging Single Shots (VSV-G) consist of pre-aliquoted, lyophilized, single tubes of Xfect Transfection Reagent premixed with an optimized formulation of VSV-G pseudotyped Lenti-X lentiviral packaging plasmids. High-titer virus is produced by simply reconstituting this mixture with your lentiviral vector of choice in sterile water and adding it to 293T cells, e.g., Lenti-X 293T Cells (Cat. No. 632180), in a 10 cm dish. Documents Components You May Also Like Image Data Back The Lenti-X Packaging Single Shots (VSV-G) protocol The Lenti-X Packaging Single Shots (VSV-G) protocol. Back Consistent, high-efficiency transfections lead to high titers Consistent, high-efficiency transfections lead to high titers. A lentiviral vector containing the ZsGreen1 gene was packaged according to the Lenti-X single shots protocol in four independent experiments. Briefly, 7 µg of pLVX-ZsGreen1 plasmid was added to each of four Lenti-X single shots; the tubes were vortexed for 20 sec and incubated at room temperature for 10 min. Then, the mixture was added to cultured Lenti-X 293T cells that were approximately 80% confluent. 48 hours after transfection, the cells were imaged by fluorescence microscopy (Panel A, top) and light microscopy (Panel A, bottom). After images were taken, the supernatant was harvested and used infect HT1080 cells for titer determination (Panel B, IFU/ml). Back High-titer virus was produced regardless of the lentiviral vector backbone with Lenti-X packaging single shots High-titer virus was produced regardless of the lentiviral vector backbone with Lenti-X packaging single shots. A CMV ZsGreen1 expression cassette was cloned into several lentiviral vector backbones. These vectors were then packaged into lentivirus using the Lenti-X packaging single shots following the provided protocol. Briefly, 7 µg of pLVX-ZsGreen1 plasmid was added to each of four Lenti-X single shots, and the tubes were vortexed for 20 sec and incubated at room temperature for 10 min. Then, the mixture was added to cultured Lenti-X 293T cells that were approximately 80% confluent. After 48 hours, titer was determined using several methods. To determine infectivity, the supernatant was harvested and used to infect HT1080 cells (Flow Cytometry). Harvested viral supernatants were also analyzed by RT-PCR to quantify viral genome copies (qRT-PCR, Lenti-X qRT-PCR Titration Kit), ELISA to measure p24 (p24 ELISA, Lenti-X p24 Rapid Titer Kit), and by a rapid lentiviral detection method (Lenti-X GoStix). Back A comparison of fourth- and third-generation lentiviral packaging systems A comparison of fourth- and third-generation lentiviral packaging systems . Our Lenti-X Packaging single shots utilize a packaging system that consists of five separate components (Panel A), mixed in proprietary proportions for optimized packaging activity. The separation of the gag, pol, and env genes effectively reduces the incidence of RCL (Wu et al., 2000). High levels of expression of essential viral components are driven by the Tet-Off and Tat transactivators, which induce a cascade of expression that results in high titers of lentivirus. The pol gene is fused to vpr to ensure transport of the reverse transcriptase/integrase protein into the recombinant lentiviral particle. Not all vector elements are shown. Other 3rd generation lentiviral packaging systems (Panel B) generate lower titers, do not contain separate gag and pol sequences, and do not use a transactivation cascade mechanism.

This lentiviral expression vector encodes an mCherry fluorescent protein tag. This bright red fluorescent protein was derived by site-directed mutagenesis of mRFP1, a monomeric mutant of DsRed. Inserting a cDNA in the MCS downstream of the mCherry coding sequence joins your protein of interest to the C-terminus of the tag, and allows the fusion protein to be tracked and studied in transduced cells. To package the vector into high-titer, replication-incompetent lentivirus, we recommend using Lenti-X Packaging Single Shots and the Lenti-X 293T Cell Line. The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

This lentiviral expression vector encodes an mCherry fluorescent protein tag. This bright red fluorescent protein was derived by site-directed mutagenesis of mRFP1, a monomeric mutant of DsRed. Inserting a cDNA in the MCS upstream of the mCherry coding sequence joins your protein of interest to the N-terminus of the tag, and allows the fusion protein to be tracked and studied in transduced cells. To package the vector into high-titer, replication-incompetent lentivirus, we recommend using Lenti-X Packaging Single Shots and the Lenti-X 293T Cell Line. The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

pLVX-PAmCherry-C1 Vector is an HIV-1-based, lentiviral expression vector. Lentiviral particles derived from the vector allow you to infect cells and express your gene of interest fused to PAmCherry, a photoactivatable mutant of the fluorescent protein mCherry. PAmCherry is non-fluorescent until photoactivated by a short exposure to light at a wavelength between 350 nm and 400 nm. The excitation/emission wavelengths of photoactivated PAmCherry are 564 nm and 595 nm. Genes cloned into the MCS will be expressed as fusions to the C-terminus of PAmCherry if they are in the same reading frame as PAmCherry and there are no intervening stop codons.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

pLVX-PAmCherry-N1 Vector is an HIV-1-based, lentiviral expression vector. Lentiviral particles derived from the vector allow you to infect cells and express your gene of interest fused to PAmCherry, a photoactivatable mutant of the fluorescent protein mCherry. PAmCherry is non-fluorescence until photoactivated by a short exposure to light at a wavelength between 350 nm and 400 nm. The excitation/emission wavelengths of photoactivated PAmCherry are 564 nm and 595 nm. Genes cloned into the MCS will be expressed as fusions to the N-terminus of PAmCherry if they are in the same reading frame as PAmCherry and there are no intervening stop codons.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

This lentiviral expression vector encodes the tdTomato fluorescent protein tag. This very bright red fluorescent protein is a genetic fusion of two copies of dTomato, which was specifically designed for low aggregation. It adopts an intramolecular tandem dimer structure that contributes to its exceptional brightness, yet it behaves like a monomer. Inserting a cDNA in the MCS downstream of the tdTomato coding sequence joins your protein of interest to the C-terminus of the tag, and allows the fusion protein to be tracked and studied in transduced cells.

To package the vector into high-titer, replication-incompetent lentivirus, we recommend using Lenti-X Packaging Single Shots and the Lenti-X 293T Cell Line. The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

This lentiviral expression vector encodes the tdTomato fluorescent protein tag. This very bright red fluorescent protein is a genetic fusion of two copies of dTomato, which was specifically designed for low aggregation. It adopts an intramolecular tandem dimer structure that contributes to its exceptional brightness, yet it behaves like a monomer. Inserting a cDNA in the MCS upstream of the tdTomato coding sequence joins your protein of interest to the N-terminus of the tag, and allows the fusion protein to be tracked and studied in transduced cells.

To package the vector into high-titer, replication-incompetent lentivirus, we recommend using Lenti-X Packaging Single Shots and the Lenti-X 293T Cell Line. The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

This lentiviral expression vector encodes the ZsGreen1 fluorescent protein tag. The coding sequence for this very bright green fluorescent protein (2.5x brighter than EGFP) was modified to include human codon usage preferences, which allow improved expression in mammalian cells. Inserting a cDNA in the MCS downstream of the ZsGreen1 coding sequence joins your protein of interest to the C-terminus of the tag, and allows the fusion protein to be tracked and studied in transduced cells. To package the vector into high-titer, replication-incompetent lentivirus, we recommend using Lenti-X Packaging Single Shots and the Lenti-X 293T Cell Line. The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

This lentiviral expression vector encodes the ZsGreen1 fluorescent protein tag. The coding sequence for this very bright green fluorescent protein (2.5x brighter than EGFP) was modified to include human codon usage preferences, which allow improved expression in mammalian cells. Inserting a cDNA in the MCS upstream of the ZsGreen1 coding sequence joins your protein of interest to the N-terminus of the tag, and allows the fusion protein to be tracked and studied in transduced cells.

To package the vector into high-titer, replication-incompetent lentivirus, we recommend using Lenti-X Packaging Single Shots and the Lenti-X 293T Cell Line. The resulting lentivirus can then be used to transduce virtually any mammalian cell type.

Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

Components

Lentivirus

Lentiviral vectors with fluorescent proteins

DsRed-Monomer & AcGFP1

ZsGreen1 & tdTomato

mCherry

DsRed-Express2

Notice to purchaser

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Overview

More Information

Lentiviral vectors with fluorescent proteins

DsRed-Monomer & AcGFP1

ZsGreen1 & tdTomato

mCherry

DsRed-Express2

Notice to purchaser