If I have samples (tails) of unknown kind of mice
but the mice should be either transgenic (heterozygous) or not (wild type
I am now have a forward primer and 2 reverse primer one mutant reverse and the other wild type reverse
CTG AAC TTG TGG CCG TTT AC Mutant Reverse
GTC TGG TCA GAA TGC CCT TG Wild type
CTG CTC TCT GCT CCC AGT CT Common
Now when I prepare the master mix to detect the kind of mice heterozygous or wild type
should I make a 2 tubes from each sample one for the common and the mutant reverse and the other with common and wild type reverse.
and see the results
or I will put the 2 reverse and one common all together
I think it doesnt make sense as how the 2 reverse work together
My expected results as follow
Mutant = 119 bp Heterozygote = 119 bp and 386 bp Wild type = 386 bp
Thanks in advance
Usually, you put one reverse and two forward primers in the same mix. You will either have one (WT or Mutant) or two bands (Heterozygote) on your gel.
For you 1st experiment, on your DNA gel, it will be a good idea to use PCR products from a known wt, mutant & het. mice as control (something that was used previously on a gel and gave the real bands)
Thanks for answer
I think you mean 1 forward and 2 reverse
As I have 2 different reverse for each kind
And the forward is the common primer