Plasmid Primer PCR product - Fraunhofer IME · 2021. 2. 25. · Contact:...
3
Contact: [email protected] Dr. Jost Muth +49 (0)241 6085 12050 Lena Freund +49 (0)241 6085 35045 Sequencing Service Fraunhofer-Institut für Molekularbiologie und Angewandte Oekologie IME Forckenbeckstr. 6 | 52074 Aachen 1 | 3 DNA Sequencing at IME Template Quality: α Template Quantity: Sample preparation: Do not use TE buffer since it contains EDTA which will inhibit the sequencing reaction. Primer 2 μL of 10 μM primer for standard plasmid & PCR product 2 - 5 μL of 10 μM primer for all other cases PCR product 100 - 200 bp 15 - 30 ng 200 - 500 bp 25 - 50 ng 500 - 1000 bp 40 - 100 ng >2000 bp 80 - 200 ng Plasmid concentration ≥100 ng/μL "standard" plasmid 80-160 ng/kb "gateway" plasmid 140-240 ng/kb Cosmid, λDNA 2-5 μL of DNA [1μg/μL] BAC, YAC, PAC 2-5 μL of DNA [1μg/μL] siRNA plasmid 140-240 ng/kb
Transcript of Plasmid Primer PCR product - Fraunhofer IME · 2021. 2. 25. · Contact:...
Contact:
Dr. Jost Muth +49 (0)241 6085 12050
Lena Freund +49 (0)241 6085 35045
Sequencing Service
Fraunhofer-Institut für Molekularbiologie
und Angewandte Oekologie IME
Forckenbeckstr. 6 | 52074 Aachen
1 | 3
DNA Sequencing at IME
Template Quality:
α
Template Quantity:
Sample preparation:
Do not use TE buffer since it contains EDTA which will inhibit the sequencing reaction.
Primer
2 µL of 10 µM primer for standard plasmid & PCR product
2 - 5 µL of 10 µM primer for all other cases
PCR product
100 - 200 bp 15 - 30 ng
200 - 500 bp 25 - 50 ng
500 - 1000 bp 40 - 100 ng
>2000 bp 80 - 200 ng
Plasmidconcentration ≥100 ng/µL
"standard" plasmid 80-160 ng/kb
"gateway" plasmid 140-240 ng/kb
Cosmid, λDNA 2-5 µL of DNA [1µg/µL]
BAC, YAC, PAC 2-5 µL of DNA [1µg/µL]
siRNA plasmid 140-240 ng/kb
Contact:
Dr. Jost Muth +49 (0)241 6085 12050
Lena Freund +49 (0)241 6085 35045
Sequencing Service
Fraunhofer-Institut für Molekularbiologie
und Angewandte Oekologie IME
Forckenbeckstr. 6 | 52074 Aachen
2 | 3
consecutive
Sample delivery:
10:30 am
8:30 am
Sequencing Order Sheet:
Sample Initial:
Template (14 µL):
Primer (2 µL, 10 pmol/µL):Tm Oligo [C°]:
Product length:PCR Product: Relevant Feature:
Plasmid Backbone & Size:
Do not forget to fill out the cells containing your name and contact information!
Contact:
Dr. Jost Muth +49 (0)241 6085 12050
Lena Freund +49 (0)241 6085 35045
Sequencing Service
Fraunhofer-Institut für Molekularbiologie
und Angewandte Oekologie IME
Forckenbeckstr. 6 | 52074 Aachen
3 | 3
Further Information:
α
The six golden rules for sequencing at IME:
A
overall GC rich
B
partial GC rich
C
GT rich region
D
homo polymer region
E
repetitive DNA
F
siRNA construct
G
gateway tech-
nology
H
large DNA
construct (>12kb)
I
BAC, YAC, PAC
J
Cosmid, Lambda
DNA
