pLenti CMV GFP Hygro (656-4)
(Plasmid #17446)

• Purpose: 3rd gen lentiviral eGFP expression vector, CMV promoter, Hygro
• Depositing Labs: Eric Campeau, Paul Kaufman
• Publication: Campeau et al PLoS One. 2009 Aug 6;4(8):e6529.

Backbone

• Vector backbone: p156RRL-sinPPT-CMV-GFP-PRE/Nhe I
  (Search Vector Database)
• Backbone size w/o insert (bp): 8591
• Vector type: Mammalian Expression, Lentiviral
• Selectable markers: Hygromycin

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin
• Growth Temperature: 37°C
• Growth Strain(s): Stbl3
• Growth instructions: Stbl3, 37oC
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: GFP
• Alt name: enhanced GFP
• Insert Size (bp): 719

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: BamHI (not destroyed)
• 3′ cloning site: Sal I (not destroyed)
• 5′ sequencing primer: CMVforw

Resource Information

• Addgene Notes:
  • Addgene diagnostic digest
  • Addgene Diagnostic Digest
• Terms and Licenses:
  • UBMTA
  • Institut Pasteur Label License for cPPT
• Industry Terms:
  • Not Available to Industry
• Articles Citing this Plasmid:
  • 10 References

Information for Lentiviral Prep (Catalog # 17446-LV)

Purpose

Ready-to-use Lentiviral Prep particles produced from pLenti CMV GFP Hygro (656-4) (#17446). In addition to the viral particles, you will also receive purified pLenti CMV GFP Hygro (656-4) plasmid DNA.

Lentiviral particles carrying the GFP and hygromycin resistance.

Delivery

• Volume: 1mL
• Titer: ≥5x10⁶ TU/mL
• Pricing: $100 USD for preparation of 1mL virus + $30 USD for plasmid.
• Storage: Store at -80℃. Thaw just before use and keep on ice.
• Shipment: Viral particles are shipped frozen on dry ice. Plasmid DNA (≥ 200ng) will also be included in the shipment.

Viral Production & Use

• Packaging Plasmids: psPAX2 (plasmid #12260)
• Envelope: pMD2.G (plasmid #12259)
• Buffer: OptiPro
• Selectable Marker: Hygromycin

Biosafety

Requestor is responsible for compliance with their institution's biosafety regulations. Lentivirus is generally considered BSL-2. AAV is generally considered BSL-1, but may require BSL-2 handling depending on the insert. Biosafety Guide

Resource Information

• Terms and Licenses:
  • Terms of Use for Viral Vectors
• Industry Terms:
  • Not Available to Industry

Viral Quality Control

Titering Method:

• ddPCR assay: 293T cells were transduced with serial dilutions of 17446-LV, harvested several days later, and genomic DNA was isolated. Copies of RRE were measured and normalized to RPP30.

Notes:

• PCR confirmation of insert: PCR was carried out with primers targeting the hygromycin resistance gene and the SV40 polyA. The PCR product was visualized on an agarose gel for size confirmation.
  Forward Primer: Hygro-internal-F GCGCCGATGGTTTCTACAAA
  Reverse Primer: SV40pA-R GAAATTTGTGATGCTATTGC

Visit our viral production page for more information.

Information for Concentrated Lentiviral Prep (Catalog # 17446-LVC)

Item is discontinued.

Addgene no longer distributes this item. Contact for more information.

Viral service discontinued.

Purpose

Ready-to-use Concentrated Lentiviral Prep particles produced from pLenti CMV GFP Hygro (656-4) (#17446). In addition to the viral particles, you will also receive purified pLenti CMV GFP Hygro (656-4) plasmid DNA.

Concentrated lentiviral particles carrying the GFP and hygromcyin resistance.

Delivery

• Volume: .
• Titer: ≥1×10⁷ TU/mL
• Pricing: $150 USD for preparation of . virus + $30 USD for plasmid.
• Storage: Store at -80℃. Thaw just before use and keep on ice.
• Shipment: Viral particles are shipped frozen on dry ice. Plasmid DNA (≥ 200ng) will also be included in the shipment.

Viral Production & Use

• Packaging Plasmids: psPAX2 (plasmid #12260)
• Envelope: pMD2.G (plasmid #12259)
• Buffer: Opti-Pro SFM
• Selectable Marker: Hygromycin
• Purification: Lentivirus was harvested from cell culture medium (DMEM + 10% FBS). Lentiviral particles were then collected by precipitation in polyethylene glycol (PEG) followed by centrifugation. Precipitated pellets containing viral particles were resuspended in PBS.

Biosafety

Requestor is responsible for compliance with their institution's biosafety regulations. Lentivirus is generally considered BSL-2. AAV is generally considered BSL-1, but may require BSL-2 handling depending on the insert. Biosafety Guide

Resource Information

• Terms and Licenses:
  • Terms of Use for Viral Vectors
• Industry Terms:
  • Not Available to Industry

Viral Quality Control

Titering Method:

• ddPCR assay: 293T cells were transduced with serial dilutions of 17446-LVC, harvested several days later, and genomic DNA was isolated. Copies of RRE were measured and normalized to RPP30.

Notes:

• PCR confirmation of insert: PCR was carried out with primers targeting the hygromycin resistance gene and the SV40 polyA. The PCR product was visualized on an agarose gel for size confirmation.
  Forward Primer: Hygro-internal-F GCGCCGATGGTTTCTACAAA
  Reverse Primer: SV40pA-R GAAATTTGTGATGCTATTGC

Visit our viral production page for more information.