On Apr 24, 11:19pm, William Jia wrote:
> Subject: PCR diagnositic technology
> Hi,
> I wonder if anybody knows whether a PCR based diagnostic technology is
> availabe for screening large number of samples in a reasonable time (say 10
> hours)? Please email to this account. Thanks in advance.
>> William
>> -- End of excerpt from William Jia <wjia at unixg.ubc.ca>
One method is to use a square grid setup. This only works if you have good
sensitivity and expect a low positivity rate. The basic method is as follows:
. . . . . . . . . . a
. . . . . . . . . . b
. . . . . . . . . . c
. . . . . . . . . . d
. . . . . . . . . . e
. . . . . * . . . . f
. . . . . . . . . . g
. . . . . . . . . . h
. . . . . . . . . . i
. . . . . . . . . . j
1 2 3 4 5 6 7 8 9 10
Assume each dot is a sample. Run twenty PCR reactions mixing all the samples
in each row (a, b, etc.) and each column (1, 2, etc.). If you know you can
detect your positive when diluted 1:10, then it's fine. All you need to do is
then determine which positive rows and columns match up and you've found your
positive sample.
In the above example, the asterisk is a positive, so in this case only row f
and column 6 would yield positive results.
In this case, you would perform 20 PCRs instead of 100 and only need to
comfirm the positive. Keep in mind that this only works if you expect few
positives.
I have seen this method in action to isolate YAC clones from a chromosome
library. In that case, grid 19 X 19 were used (38 PCRs for screening 361
different clones) for about a 10 fold efficiency.
It seems that once the sample number gets large, the hands on time is greatly
increased. This method reduces the number of PCRs to screen a large sample.
Mike Kurilla
