cells. pRPCX1.0 is a Moloney murine leukemia virus derived retroviral gene delivery and
expression vector. Expression of inserted cDNAs is powered by human cytomegalovirus immediate
early gene promoter, ensuring a high level of expression. Integration of this vector into host
genome can be selected for resistance to puromycin. The finished cDNA constructs can be used
directly as plasmids in transfection experiments, or packaged into virus particles in retroviral
packaging system for infection of proliferating cells.
To order this service, please download "cDNA cloning order form" (PDF, Word format) and send
your starting clone along with it to
Cellogenetics, Inc
10075 Tyler Place, Suite 6
Ijamsville, MD 21754
If you don't already have the cDNA, we also offer a service to identify and isolate a full length
clone from EST libraries. Please add this service in the order form.
If you like to mark the cDNA, you may do so in the order form. Please specify the desired tag (HA,
Myc, Flag, or other common tags) and its position (5' or 3').
We also offer to modify the cDNA for making dominant negative or constitutive active , or other
desired mutants. Please specify the desired sequence change(s) in the order form.
5'-LTR: 1-589
Packaging signal: 659-1468
Puromycin resistance: 1512-2306
CMV promoter: 2656-3393
Multiple cloning site: 2949-3066
ColE1 origin: 3991
b-lactamase gene: 5611-4751
5' Sequencing primer: 2863-2884
5'-GTCAGATCGCCTGGAGACGCC-3'
3' Sequencing primer: 3143-3122
5'-TAGCTTGCCAAACCTACAGGT-3'
Suitable host strain: DH5a, Stbl-2 and others
E. coli replication origin: ColE1
Antibiotics resistance: Ampicillin (100 ug/ml)
Viral packaging cells: Pheonix A or Pheonix E cells
Reference:
1. Miller A.D. & Buttimore C. (1986) Mol Cell Biol.
6:2895-902
2. Miller A.D. & Chen F. (1996)J Virol. 70:5564-71
3. Choy L. et al (2000) JCB 149:667-681
Purchase of the cDNA subcloning service amounts to entering a service agreement under which
Cellogenetics is contractually obligated to:
1. design and prepare DNA oligo primers,
2. perform high fidelity PCR reaction to amplify cDNA with appropriate modification,
3. subclone the PCR fragment into the pRPCX1.0 vector,
4. prepare plasmid DNA and verify cDNA insert by restriction enzyme digestion,
5. sequence both ends of the finished clone to confirm correct cloning junctions and tags,
6. ship out the finished constructs as glycerol stocks.
