Custom Lentivirus Constructs

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Please view our ordering information for custom constructs to learn how to register for an account, place orders for in-stock vectors and make payment through our online system.

Pricing

New Custom Vectors: $1,342 for 500ul of concentrated virus. Re-order Custom Vectors: $1,342 for 500ul of concentrated virus.

Feline Immunodeficiency Virus (FIV) Vectors

The FIV based vector system is generated by triple plasmid transfection.

1. The packaging plasmid encodes the necessary viral proteins in trans, but it is not packaged into the particles. Thus, the vector is non-replicating.

2. The transgene plasmid contains the expression cassette, comprised of the gene and promoter of interest. It also retains the minimal cis acting viral sequences necessary for packaging, reverse transcription and integration.

3. The envelope plasmid provides for the viral envelope protein. Typically, the envelope is VSV-G that has broad tropism and proven packaging efficiency; however, other envelopes can be substituted to alter tropism via a process called pseudotyping.

Material to be provided by the investigator: The gene interest driven by the appropriate promoter must be cloned into the transgene plasmid. A variety of transgene shuttle plasmids [link to shuttle page] with multiple cloning sites are available for this purpose. These plasmids are owned by the Chiron/Novartis Corporation and are provided to the investigator after a Material Transfer Agreement is signed.

For new custom FIV vector production, the investigator will need to provide at least 250 μg of transgene plasmid expressing the gene of interest at a concentration of at least 0.25 μg/μl. We require a map and sequence along with a picture of a confirmatory restriction digest of the transgene plasmid to the Viral Vector Core prior to production.

Timeline: 3-4weeks from the time the transgene plasmid is received.

Services include:

  •  Transfection and supernatant collection.
  • Ultracentrifugation concentration.
  • Determination of infectious titer (TU/ml).
  • 500 μl of concentrated virus is provided at a titer no less than 1x107TU/ml but potentially greater than 5x108 TU/ml.
  • $1,342 for 500ul of purified vector in 100ul aliquots.

Vehicle: Unless otherwise directed, lentiviral vectors are re-suspended in a standard 4% Lactose/PBS buffer for stability.

References

Johnston JC, Gasmi M, Lim LE, Elder JH, Yee JK, Jolly DJ, Campbell KP, Davidson BL, Sauter SL. Minimum requirements for efficient transduction of dividing and nondividing cells by feline immunodeficiency virus vectors. J Virol. 1999; 73:4991-5000 

Sinn PL, Goreham-Voss JD, Arias AC, Hickey MA, Maury W, Chikkanna-Gowda CP, McCray PB Jr. Enhanced gene expression conferred by stepwise modification of a nonprimate lentiviral vector. Hum Gene Ther. 2007; 18:1244-52.

Human Immunodeficiency Virus (HIV) Vectors

Third Generation HIV

Invitrogen Corporation ViraPowerTM Lentivirus Expression System (Cat# K4975-00). The third generation HIV-based vector is generated by four plasmid transfection.

1. The packaging plasmid encodes the necessary viral proteins in trans, but it is not packaged into the particles. Thus, the vector is non-replicating.

2. The transgene plasmid contains the expression cassette, comprised of the gene and promoter of interest. It also retains the minimal cis acting viral sequences necessary for packaging, reverse transcription and integration.

3. The envelope plasmid provides for the viral envelope protein. Typically, the envelope is VSV-G that has broad tropism and proven packaging efficiency; however, other envelopes can be substituted to alter tropism via a process called pseudotyping.

4.The Rev protein is provided by an independent expression plasmid.

Material to be provided by the investigator: The gene of interest driven by the appropriate promoter must be cloned into the transgene plasmid. The investigator will need to purchase the transgene shuttle plasmid from Invitrogen or provide a compatible plasmid. The Viral Vector Core is not licensed to distribute these plasmids.

For new custom HIV vector production, the investigator will need to provide at least 250 μg of transgene plasmid expressing the gene of interest at a concentration of at least 0.25 μg/μl. We require a map and sequence along with a picture of a confirmatory restriction digest of the transgene plasmid to the Viral Vector Core prior to production.

Second Generation HIV

Open Biosystems Trans-LentiviralTM Packaging System (Cat# TLP4691). Note: the second generation system can package the third generation transgene plasmid, but not vice, versa. The second generation HIV-based vector is generated by three plasmid transfection.

1. The packaging plasmid encodes the necessary viral proteins in trans, but it is not packaged into the particles. Thus, the vector is non-replicating.

2. The transgene plasmid contains the expression cassette, comprised of the gene and promoter of interest. It also retains the minimal cis acting viral sequences necessary for packaging, reverse transcription and integration.

3. The envelope plasmid provides for the viral envelope protein. Typically, the envelope is VSV-G that has broad tropism and proven packaging efficiency; however, other envelopes can be substituted to alter tropism via a process called pseudotyping.

Material to be provided by the investigator: The gene interest driven by the appropriate promoter must be cloned into the transgene plasmid. The investigator will need to purchase the transgene shuttle plasmid from Open Biosystems. The Viral Vector Core is not licensed to distribute these plasmids.

For new custom HIV vector production, the investigator will need to provide at least 250 μg of transgene plasmid expressing the gene of interest at a concentration of at least 0.25 μg/μl. We require a map and sequence along with a picture of a confirmatory restriction digest of the transgene plasmid to the Viral Vector Core prior to production.

Timeline: 3-4 weeks from the time the plasmid is received.

Services include:

  • Transfection and supernatant collection.
  • Ultracentrifugation concentration.
  • Determination of infectious titer (TU/ml).
  • 500 μl of concentrated virus is provided at a titer no less than 1x107TU/ml but potentially greater than 5x108 TU/ml.
  • $1342 for 500ul of purified vector in 100ul aliquots.
  • Vehicle: Unless otherwise directed, lentiviral vectors are re-suspended in a standard 4% Lactose/PBS buffer for stability.

ViraPowerTM Lentivirus Expression System References:

LinksZufferey R, Dull T, Mandel RJ, Bukovsky A, Quiroz D, Naldini L, Trono D.Self-inactivating lentivirus vector for safe and efficient in vivo gene delivery. J Virol. 1998 Dec;72(12):9873-80.

LinksDull T, Zufferey R, Kelly M, Mandel RJ, Nguyen M, Trono D, Naldini L. A third-generation lentivirus vector with a conditional packaging system. J Virol. 1998 Nov;72(11):8463-71

Trans-LentiviralTM Packaging System References:

Kappes & Wu, Somatic Cell and Molecular Genetics, Vol. 26, Nos. 1/6, November 2001.

Kappes JC, Wu X, Wakefield JK. Production of trans-lentiviral vector with predictable safety. Methods Mol Med. 2003; 76:449-65.

Kappes JC, Wu X. Safety considerations in vector development. Somat Cell Mol Genet. 2001 Nov;26(1-6):147-58

Liu H, Wu X, Xiao H, et al. Incorporation of functional human immunodeficiency virus type 1 integrase into virions independent of the Gag-Pol precursor protein. J Virol 1997; 71:7701-7710.

Moloney Murine Leukemia Retrovirus (mMLV) Vectors

mMLV-based vectors are generated by triple transfection.

1. The packaging plasmid encodes the necessary viral proteins in trans, but it is not packaged into the particles. Thus, the vector is non-replicating.

2. The transgene plasmid contains the expression cassette, comprised of the gene and promoter of interest. It also retains the minimal cis acting viral sequences necessary for packaging, reverse transcription and integration.

3. The envelope plasmid provides for the viral envelope protein. Typically, the envelope is VSV-G that has broad tropism and proven packaging efficiency; however, other envelopes can be substituted to alter tropism via a process called pseudotyping.

Material to be provided by the investigator: The gene interest driven by the appropriate promoter must be cloned into an appropriate transgene plasmid. The investigator will need to provide their own transgene shuttle plasmid because the Viral Vector Core is not licensed to distribute these plasmids.

For new custom mMLV vector production, the investigator will need to provide at least 250 μg of transgene plasmid expressing the gene of interest at a concentration of at least 0.25 μg/μl. We require a map and sequence along with a picture of a confirmatory restriction digest of the transgene plasmid to the Viral Vector Core prior to production.

Timeline: 3-4 weeks from the time the plasmid is received.

Services include:

  • Transfection and supernatant collection.
  • Ultracentrifugation concentration.
  • Determination of infectious titer (TU/ml).
  • 500 μl of concentrated virus is provided at a titer no less than 1x106TU/ml but potentially greater than 5x107 TU/ml.
  • $1342 for 500ul of vector in 100ul aliquots. 

Vehicle: Unless otherwise directed, retroviral vectors are re-suspended in a standard 4% Lactose/PBS buffer for stability.