pSLIC-PGfpP2A^TM

AbDia Technologies Inc. provides pSLIC-GfpP2A lentivirus plasmid construction kit (LT2003) to let user creates the lentivirus plasmid for the gene of interest. This kit comes with SLIC enzyme and a lentivirus plasmid which contains GFP expression sequence and P2A sequence.

SLIC (sequence and ligation independent cloning) allows users to insert PCR product into the plasmid without ligation. P2A peptide sequence allows to express both GFP gene and user’s gene of interest by a single CMV promoter (bicistronic). P2A sequence in our plasmid is GSGATNFSLLKQAGDVEENPG/PD. The first part (before slash "/ ") GSGATNFSLLKQAGDVEENPG is attached to GFP and the second part (after slash "/ ") PD is attached to the gene of interest users insert. Our proprietary SLIC enzyme system which has superb performance and temperature stability (stable even at room temperature for a few days). In 30min reaction with SLIC enzyme, PCR product is inserted downstream of P2A sequence.

This kit is suitable to make lentivirus for primary cells in which antibiotic selection is difficult to perform but monitoring expression rate is important. In most of the cells it takes 1-3 days to express GFP and target proteins. Because of bicistronic expression, GFP positive cells are also positive for a gene of interest expression.

Components and Storage

Catalogue #	Component	Size	Storage
LT2003-01	pSLIC-GfpP2ATM Plasmid	20µl (200ng/µl)	4°C.This plasmid is not intended to amplify.
SE1001-01	10×SLIC-EnzymeTM	25µl	-20°C, Enzyme is stable at least 4 days at room temperature.
Puro1001-01	GFP Template DNA (control)	10µl (10ng/µl)	4°C
Puro1002-01	GFP Forward and Reverse Primer Mixture	10µl, 10µM	4°C

The kit contains enough plasmids and reagents to perform at least 20 reactions for creating lentiviral vector plasmids which contain gene of interest.

Kit Components and Storage Conditions

Catalogue #	Component	Size	Storage Condition
LT1001-1	Lentivirus production medium	55mL	4 °C. Add 1.2ml FBS before use.
LT1001-2	Helper-plasmids mixture	0.2mL	4 °C
LT1001-3	Production helper-solution	0.5mL	4 °C
LT1001-4	Plate coating solution	2.0mL	4 °C
LT1001-5	Lentivirus Purification Solution	20mL	4 °C
LT1001-6	Lentivirus storage solution	2.0mL	4 °C
LT1001-7	GFP expressing lentivirus plasmid	1µg in 20µl TE	4 °C. Ampicillin resistance

References

1. Y. Ido, et al. PLoS ONE, 2012 Apr 07(4): e35092


2. Mamie Z Li & Stephen J Elledge Nature Methods 2007; 4:251-256


3. Gibson DG Methods Enzymol 2011; 498:349-61


4. Jin Hee Kim, Sang-Rok Lee, Li-Hua Li, et al. PLoS ONE 2011 Apr 06 (4): e18556


5. Georgios Trichas, Jo Begbie and Shankar Srinivas BMC Biology 2008, 6:40