Hi, how much cdna you are taking for PCR? You can try taking more amount of cdna. Also increase the number of cycles in PCR.

Thank you Debaleena Kar. I used 2 µl of cDNA and 25 cycle in PCR

Hi Pravin,

it depends on how is transcribed the GAPDH gene in the brain in respect to the average transcription and the amount of cDNA you used. for this point, it's preferred to use ng with which you can figure the amount is needed to increase the chance to detect the gene. increasing cycles will gave nothing if the gene is not expressed in brain....one other important thing is the quality of the designed primers and the right protocol for PCR you're using.

conditions of the individuals are also really important, see Article Oxidatively Modified Glyceraldehyde-3-Phosphate Dehydrogenas...

fred

Thank you Frederic Lepretre

Dear Pravin...

There are many possibilities, but you may pay attention to the following problems:

• Use the right percentage of agarose for your desired size range (small products will be fuzzier when you are using a lower gel percentage).
• There are many different grades and qualities of agarose.  Some are specific for resolving certain size ranges with precision.
• The bands can slowly diffuse if it sit for some time after running.
• Load the right size of PCR product into the well. You may even loading more DNA and/or use narrower wells which will not allow the DNA to spread out in the well and the bands will be thicker.