What is pLKO plasmid?

pLKO. 1 is a replication-incompetent lentiviral vector chosen by the TRC for expression of shRNAs. pLKO. 1 can be introduced into cells via direct transfection, or can be converted into lentiviral particles for subsequent infection of a target cell line. Once introduced, the puromycin resistance marker encoded in pLKO.

What do packaging plasmids do?

The envelope plasmids and packaging plasmids provide all of the proteins essential for transcription and packaging of an RNA copy of the expression construct into recombinant pseudoviral particles.

Can lentiviral plasmids be used in direct transfections as opposed to making virus?

Can lentiviral plasmids be used in direct transfections as opposed to making virus? Some (but not all) lentiviral transfer plasmids can be used in transient transfections to achieve expression of the transgene, and those that can are primarily third generation constructs.

What is pcdna3?

This pcDNA™3.1(-) vector is designed for high-level, constitutive expression in a variety of mammalian cell lines. It contains a Geneticin™ selectable marker and a reverse-orientation multiple cloning site. The pcDNA™3.1 Expression Vector Family.

What is meant by lentivirus?

Definition of lentivirus : any of a genus (Lentivirus) of retroviruses (such as the HIVs and SIV) that cause slowly progressive often fatal human and animal diseases (such as AIDS)

What is Wpre sequence?

Woodchuck Hepatitis Virus (WHP) Posttranscriptional Regulatory Element (WPRE) is a DNA sequence that, when transcribed creates a tertiary structure enhancing expression. Commonly used in molecular biology to increase expression of genes delivered by viral vectors.

What is viral packaging?

Genome packaging is a fundamental process in a viral life cycle. Many viruses assemble preformed capsids into which the genomic material is subsequently packaged. These viruses use a packaging motor protein that is driven by the hydrolysis of ATP to condense the nucleic acids into a confined space.

How do lentiviruses work?

Lentiviruses (a genus of retrovirus) express reverse transcriptase, which converts the viral RNA to double stranded DNA, and integrase, which inserts this viral DNA into the host DNA. Once the viral DNA is integrated into the host DNA, it divides along with host cell and none are the wiser.

Which plasmids are recommended for use with the plko?

The following plasmids available from Addgene are recommended for use in conjunction with the pLKO.1 TRC-cloning vector. Negative control vector containing non-hairpin insert.

What is the plko vector?

pLKO.1 is a replication-incompetent lentiviral vector chosen by the TRC for expression of shRNAs. pLKO.1 can be introduced into cells via direct transfection, or can be converted into lentiviral particles for subsequent infection of a target cell line.

What is the plko protocol used for?

This protocol is provided for your convenience. See “warranty information” in appendix. The pLKO.1 cloning vector is the backbone upon which The RNAi Consortium has built a library of shRNAs directed against 15,000 human and 15,000 mouse genes.

What is lentiviral plko?

Lentiviral plKO.1 construct coding for a short-hairpin RNA targeting the human gene DDX58 (coding for RLR sensor RIG-I). Contains a puro resistance cassette. Lentiviral plKO.1 construct coding for a short-hairpin RNA targeting the human gene TBK1. Contains a puro resistance cassette. Orimo et al Cell. 2005 May 6. 121 (3):335-48.