can anyone tell me what would be the change in the bands on gel if i use primers from exon-exon junctions and non exon-exon junction??
it depends on what your template is. If it is cdna then exon exon junction primers will work but genomic dna will not amplify any bands. If your primers are intronic then cdna will not amplify and genomic dna might amplify depending on the size of the amplimer. If the primers are exon to exon then cdna will amplify and genomic dna will either amplify a larger fragment if the exons are close together or will not amplify if the intervening intron is very large
Dear Paul
for instance, i am designing the primers for my cDNA my first choice is exon-exon junction primers but they are unavailable so i go for non exon exon junction primers, so there is any change regarding bands on gel or not??
regards
zaid
you must not use 2 primers within the same exon or you will amplify genomic dna at the same size as the cdna amplimer. If you amplify between primers in different exons you will get 2 bands...a large one from genomic dna including the intron(s) and a small one from the cDNA. If your introns are large then you might get no amplified band from genomic dna because the amplimer is too long. This is often the case when many introns lie between the primers
- Badges
- Science method