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How to select Gentarget Ins’s lentivirus products!

Gentarget provides variety of lentivirus products for the same target (cDNA, gene, marker, report, and so on), driven by different promoter and carrying different selection marker. Then, How to select the product? The following video provide a brief description.

Select the Promoter:

1. Promoter strength is the first to consider because it is cell type dependent. For example CMV promoter is the strongest promoter in most cell types, but is barely active in some stem cell type and weak in neuron cells. For those cell types, the CAG promoter was approved a strong one. Another example, the EF1a promoter is less cell or tissue sensitive in terms of promoter strength, and general weak and later in function than CMV promoter. You have to test the promoter strength if you do not know it. Otherside, select the EF1a promoter will be a safe pit. You can use the lentivirus products express a fluorescent marker (like GFP or RFP) under different promoters to test each promoter’s strength in your cell types. Gentarget have the report / marker driven by all kinds of promoters, like CMV, inducible CMV (TetCMV), EF1a, CAG, mPGK, Ubc, ActinB (ActB), pathway specific promoter, tissue specific promoter, and more.
2. Promoter silencing is the 2nd factor to consider when you select the promoter. CMV has tendency in some cell types, to be silenced by methylation in long term cell culture. So if you try to use lentivirus to generate a stable cell line and use it for long term culture, you need to know the CMV promoter does not silenced in your cell type or try to use EF1a promoter.
3. Pathway specific promoter and tissue specific promoter: In some cases, you want use a desired promoter for your tissue or cells, or a pathway specific promoter for investigating the specific signal transduction. Gentarget Provides all kinds of lentivirus products for the pathway and tissue specific applications.

Select the marker:

In order to select the positive transduced cells for long term expression, lentivirus often contains a fluorescent marker (for cell sorting) or /and an antibiotic marker (for killing selection).

1. Antibiotic Selection types: GenTarget’s products are made with different antibiotic selection marker, Blasticidin (Bsd), Puromycin (Puro), Neomycin (Neo), Zeocin (Zeo) or Hygromycin (Hygro).  Some products also contain a fluorescent marker, GFP, RFP, BFP, CFP or YFP, and sometime an luminance marker (luciferase). Depend your preference with the selection marker, you pick the product containing your desired antibiotic marker, Bsd, or Puro or Neo or Hygro.
2. Fluorescent Marker: If you need the product with a fluorescent marker as well, you pick the product with both antibiotic marker and fluorescent marker 9dual marker).
3. Each antibiotic has different killing curve in each cell type, which have to be tested prior to the antibiotic selection. Puromycin and Blasticidin has the “fast killing feature” (often in about one week). Most cell types are extremely sensitive to puromycin. So, you select the product with the antibiotic marker good for your positive cell selection..
4. Each fluorescent marker has different wavelength (Ex and Em). The fluorescent maker provides an easy transduction efficiency check (under Fluorescent-microscope) or cell selection (via FACS sorting). You pick the product having your desired fluorescent marker. All fluorescent markers in GenTarget’s products, are engineered with the strongest signal intensity.

Select the product format:

GenTarget’s lentivirus provides in three formats based upon its titer levels. You pick the virus depends on your application.

1. Regular lentivirus: provided in DMEM medium (1×10^7 Ifu/ml titer) with 10% FBS and pre-added polybrene (10x). It satisfies most application. Simply add the virus into your cell culture.
2. Concentrated Lentivirus: provided in PBS solution with higher titer (1×10^8 Ifu/mlm, premixed with 10x Polybrene). It can be used for the cell types having low transduction efficiency (hard to infected cell types), or used for serum sensitive cell culture, like stem cell, or some primary cells.
3. Ultra titer lentivirus: concentrated virus with ultra high titer (>=10⁹ IFU/ml)  provided in PBS. It best fits for in vivo manipulation for direct injection, gene-therapy research and application that requires extreme high titer lentivirus.