Drosophila Gene Collection
Please note that the clone(s) being sent have tested negative in our
T1 phage contamination assay. However these clones should still be handled
with care as no phage assay can be guaranteed to be 100% reliable.
THE DISPATCH NOTE SHOWS THE ORIGINAL CLONE ID ORDERED
AND ALSO THE ALIAS (PLATE AND WELL NUMBER ). THIS IS WHAT IS SHOWN ON
THE CLONE LABEL. PLEASE KEEP THE DISPATCH NOTE FOR FUTURE REFERENCE.
WHAT TO DO WITH YOUR CLONES WHEN YOU RECEIVE THEM
Plates- contain LB broth with 8% glycerol and antibiotic*.
are sent on Dry Ice. These should be stored at -70°C
Individual clones have been streaked onto LB agar containing
antibiotic*. Please store them at 4°C (not in a freezer).
As soon as possible, re-streak the clones onto the same medium and incubate
overnight at 37°C to obtain single colonies.
*Antibiotic
DGCr1.0
Plates 1-4 Ampicillin 75µg/ml
Plates 5-17 Chloramphenicol 25µg/ml
DGCr2.0
Plates 1-10 Ampicillin 75µg/ml
Plates 11-16 Chloramphenicol 25µg/ml
A selection of at least 10 single colonies should be picked
and grown in LB broth containing appropriate antibiotic + 8% glycerol
for subsequent freezing at -70°C. These frozen stocks can then be
re-grown for plasmid isolation and purification.
Streaking out for single colonies is essential in case of contamination
or cross-contamination, or if modifications of the original clone have
taken place as the library has been replicated. If there is a problem,
the more single colonies you streak out, the more likely it is that
you can retrieve the original construct. You can also isolate the original
clone from a contaminated sample by following this procedure.
The Drosophila Gene Collection r1.0 is the first
release of ~6,000 cDNA clones which has been produced by a consortium
of Berkeley Drosophila Genome Project (http://www.fruitfly.org).
The cDNA libraries have been produced in pBlueScript and pOT2, pOTB7
and pFLC-1 vector systems originating from a variety of fly tissues/organs.
Full list of cDNA libraries and mRNA sources can be found in Table 1.
DGCr3.0
Plates 1-3 Ampicillin 75µg/m
Plates 4-6 Chloramphenicol 25µg/ml
Table 1 : Description of cDNA libraries
µg/m
| Library code |
mRNA source |
| CK |
Rough endoplasmic reticulum (ER) from 8-16 hr embryos |
| GH ZAPII |
Adult heads from an isogenic y; cn bw sp strain |
| GH pOT2 |
Adult heads from an isogenic y; cn bw sp strain (remade) |
| GM |
Ovaries, stage 1-6 of oogenesis |
| HL |
Adult heads from an isogenic y; cn bw sp strain |
| LD |
Embryos (0-22 hr) |
| LP ZAPII |
Varying stages of larvae and early pupae from an isogenic
y; cn bw sp strain |
| LP pOT2 |
Varying stages of larvae and early pupae from an isogenic
y; cn bw sp strain |
| SD pOT2 |
Schneider L2 tissue cultured cells |
| AT pOTB7 |
Adult male testes and seminal vesicles dissected from 0-3
day old Ore-R males |
| RE pFlc1 |
0-22 hr mixed stage isogenic y; cn bw sp strain embryos |
| RH pFlc1 |
Adult heads from an isogenic y; cn bw sp strain |
The Drosophila Gene Collection releases 1.0 and
2.0 together consist of ~11,000 cDNA clones which have been produced
by a consortium of Berkeley Drosophila Genome
Project (http://www.fruitfly.org).
The cDNA libraries have been produced in pBlueScript and pOT2, pOTB7
and pFLC-1 vector systems originating from a variety of fly tissues/organs.
Vector information:
Please click on the links below to view desired vector map:
pOT2
http://www.fruitfly.org/about/methods/pOT2vector.html
pOTB7 http://www.fruitfly.org/about/methods/pOTB7vector.html
pFLC-1 http://www.fruitfly.org/about/methods/pFLC-Ivector.html
pBS http://www.stratagene.com/displayProduct.asp?productId=267
Further information- http://www.fruitfly.org/EST/faq.html#cdna-6
Sizing cDNA inserts:
Template: 3.0 µL from 1:50 dilution in water of frozen stock.
Use the following primer pairs:
pOT2 vector
PM001a: 5' GTCGACGTTAGAACGCGGCTAC 3'
PM002a: 5' GGGTTAAATTCCCGGGTACTGC 3'
BS vector
SK-30: 5' GGGTAACGCCAGGGTTTTCC 3'
SKMet: 5' ATGACCATGATTACGCCAAGC 3'
pFLC-1 vector
T7 primer: AAT ACG ACT CAC TAT AGG
T3 primer: AATTAACCCTCACTAAAGG
Sequencing of inserts:
Sequencing Primers for BS and pOTB7
M13 forward (-21) 18-mer: TGTAA AACGA CGGCC AGT
M13 reverse 17-mer: CAGGA AACAG CTATG AC
Sequencing Primers for pOT2
T7 primer: AAT ACG ACT CAC TAT AGG
PM001 primer: CGT TAG AAC GCG GCT ACA AT
Sequencing Primers for pFLC-1
T7 primer: AAT ACG ACT CAC TAT AGG
T3 primer: AATTAACCCTCACTAAAGGG
Alternatively, the following primers can also be used
FLC2: ATTGGAGCTCCCCGCGGTGG
KST3: CGCAATTAACCCTCACTAAAGG
Citation in publications:
Please cite the following paper for BDGP clones or sequence data.
Mark Stapleton*†, Joe Carlson*†, Peter Brokstein*†‡,
Charles Yu*†,
Mark Champe*†§ Reed George*†, Hannibal Guarin*†,
Brent Kronmiller*†,
Joanne Pacleb*†, Soo Park*†, Ken Wan*†, Gerald M Rubin*¥#
and
Susan E Celniker* A Drosophila full-length cDNA resource. Genome Biology,
2002;3:research0080.1-0080.8
Stapleton M, Liao G, Brokstein P, Hong L, Carninci P,
Shiraki T, Hayashizaki
Y, Champe M, Pacleb J, Wan K, Yu C, Carlson J, George R, Celniker S,
Rubin
GM. The Drosophila Gene Collection: Identification of Putative Full-Length
cDNAs for 70% of D. melanogaster Genes. Genome Research. 2002; 12:1294-1300.
Rubin GM, Hong L, Brokstein P, Evans-Holm M, Frise E, Stapleton M, and Harvey DA:
A Drosophila complementary DNA resource. Science 2000,
287:2222-2224.
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