Biology Reference
In-Depth Information
3. Preheat the PCR machine block to 95
, add the tubes, and then run the following
program:
95
1.
:30
67
2.
:30-1.0 per cycle
72
3.
:30
4.
Go to 1 12 times
95
5.
:30
55
6.
:30
72
7.
:30
8.
Go to 5 24 times
72
9.
2:00
22
10.
For ever
4. Run half of the sample for each reaction on a 4% agarose gel. If your reaction
conditions are working properly, for each primer set you should see an upper band
that is present in all lanes (arrowhead in
Fig. 2
), one lower band that is specific to
N2, and another lower band that is specific to CB4856 (arrows in
Fig. 2
). The two
lower bands may not be of equal intensity in the lysate that contains both N2 and
CB4856 worms; however, their intensities relative to each other should remain
constant in any lysate that contains equal amounts of N2 and CB4856 DNA at a
Fig. 2
An example of ARMS-PCR mapping. Results for SNPs on chromosomes I and IVare shown for
four different zygotic-effect recessive mutants, new1-new-4. new-1 is clearly linked to the SNP on
chromosome I, whereas new-3 is linked to the SNP on chromosome IV. Arrowhead indicates upper band
that is amplified from both N2 and CB4856. Arrows indicate lower products specific to N2 and CB4856.
The size markers used are pGEM
1
markers from Promega (the bands in the triplet near the N2- and
CB4856-specific bands are 222, 179, and 126 bp).
Search WWH ::
Custom Search