Improved third-generation lentiviral packaging with pLKO.1C vectors.

BioTechniques
Sunhye Lee, David Cobrinik

Abstract

A widely used third-generation lentiviral packaging system produces virus with enhanced biosafety by eliminating HIV accessory genes and separating packaging elements into three different plasmids. However, for certain vectors such as pLKO.1, third-generation safety features reduce lentiviral titers due to the lack of the accessory gene tat. Here we present a way to improve virus production and target gene knockdown with a modified pLKO.1 CMV pLKO.1C) vector and optimized packaging construct ratios. Replacing the pLKO.1 RSV promoter with the Cytomegalovirus promoter yielded an average of threefold higher titer than standard pLKO.1 packaged using the third-generation system, while optimizing the packaging vector ratios further increased titer and yielded an average of tenfold higher titer than pLKO.1 packaged with the second-generation system.

References

Nov 5, 1997·Nature Biotechnology·R ZuffereyD Trono
Sep 12, 1998·Journal of Virology·H MiyoshiI M Verma
Oct 10, 1998·Journal of Virology·T DullL Naldini
Aug 25, 2006·Nature Methods·David E RootDavid M Sabatini
Sep 26, 2014·Nature·Xiaoliang L XuDavid Cobrinik
Sep 10, 2016·Science·Luigi NaldiniInder M Verma

❮ Previous
Next ❯

Citations


❮ Previous
Next ❯

Methods Mentioned

BETA
FACS
transfection
gene knockdown

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.