orage of single cells:
Single cells may be isolated by different methods
(e.g., by flow cytometry or by micromanipulation).
Keep samples cool during the cell-isolation procedure
to prevent DNA degradation.
Transfer cell into a PCR tube that has been filled with
20 l of 1x PCR Buffer. Immediately freeze the sample
on dry ice.
Store cell at 80C until required for PCR analysis.
Prepare a fresh master mix for single-cell PCR (see
Thaw the cells on ice.
Distribute 30 l of the master mix into each PCR
tube, and place the tubes in the thermal cycler. Immediately
start the cycling program with a 10 min incubation step at
to activate HotStarTaq DNA Polymerase for single-cell
PCR. 50 cycles of PCR may be required to amplify a single-copy
gene in one round of PCR.
Master mix preparation:
Prepare a master mix that has a final volume of 30 l
per PCR, as detailed below.
Addition of carrier nucleic acid is usually required
(e.g., E. coli 5S rRNA).
Use polyacrylamide gel- or HPLC-purified primers only.
Source:Page: All 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 Related biology technology :1
. HotStarTaq DNA Polymerase2
. QIAGEN Multiplex PCR Handbook3
. TransMessenger Transfection Reagent Handbook4
. Transfection Reagent Selector Kit Handbook