| Product title:
| CHICKEN CGI LIBRARY
|
| Description:
| Chicken CpG library supplied as live cells
|
| Batch number:
| 96/1
|
| Date of production:
| July 1996
|
Contents of package:
- (a) 100µl aliquot of Chicken CGI library. (store
at -70°C)
- (b) 100µl aliquot of PCR primer 3558. (store at
-20°C)
- (c) 100µl aliquot of PCR primer 3559. (store at
-20°C)
On Arrival:
Contents should be stored at the above conditions until required.
We suggest that aliquots for freezing are prepared at the same time as
first thawing.
Specific product information:
| Titre:
| 108 cells/ml (107 cells/100µl) in
25% glycerol + 50µg/ml ampicillin in LB broth
|
| Host Strain:
| E.coli strain SURE MRF'
|
| Vector:
| pGEM-5Zf(-)
|
| Insert excision:
| NdeI
|
| Average insert size:
| 670 bp
|
| Growth conditions:
| LB + 50µg/ml ampicillin
|
| Coverage:
| ~ 5 fold
|
- Primers:
- Provided as a 10µM solution
5'-CGG CCG CCT GCA GGT CGA CCT TAA- 3' (Geneservice Ltd. no 3558)
5'-AAC GCG TTG GGA GCT CTC CCT TAA-3' (Geneservice Ltd. no 3559)
Additional primer stocks can be obtained through Geneservice Ltd.,
quoting the above reference numbers.
- Amplification of CpG island fragments:
We have found that the amplification of the CpG island fragments can
be achieved in a 20µl reaction volume in the following reaction
buffer :
GeneAmp PCR Buffer (10x): Tris-HCL pH 8.3 100mM
(Perkin-Elmer) KCL 500mM
MgCl2 15mM
Gelatin 0.01%
The following PCR cycle has been successfully used for amplification of
the CpG island fragments. We strongly recommend that the conditions are
optimized, to allow for lab to lab variation.
30x cycles of: 94º C 1 min
55º C 1 min
72º C 3 min
- Quality Control:
Our quality control work showed that the above primers with the conditions
stated amplified the CpG island fragments successfully. Alternative
primers, T7 and SP6 also carry out successful amplification of inserts.
5' -TAA TAC GAC TCA CTA TAG GG- 3' (T7) ;
5' -GAT TTA GGT GAC ACT ATA G- 3' (SP6)
-
- References:
The construction of the Chicken CGI library is described in:
CpG islands of chicken are on concentrated microchromosomes
McQueen HA, Fantes J, Cross SH, Clark VH, Archibald Al and Bird AP.
Nature Genetics, 1996 12(3), 321-324
