i would like some details like a picture of the gel, pcr conditions and type of agarose. Do you have no pcr product or a smear of products. It is very easy to over amplify on re pcr given that 10 cycles of pcr is 1000 times more product so you need to start with very little product ( less than 1ul) and few cycles. If the gel is normal agarose not a low gelling agarose then there are pcr inhibitors present so adding a bit more magnesium in the pcr mix may help and also lowering the annealing temperature 2degrees. Why are you re amplifying with the same primers...is it just to check the  reaction because you will need the GC clamp when you run the DGGE. If your original pcr product  is a small fragment then you may pick up some primer dimer in the cutting process and this will take over the reaction and all you will get is primer dimer

Maybe you should make a DNA extraction from the gel (homemade or by kit).

If you have the option of having different temperatures in your thermocycler, you could make the PCR in 4 different temperatures at the same time (near the annealing temperature), with 1.5, 2 and 3 mM of MgCl2 and add one extra tube for each temperature in the 2 mM of MgCl2 with 10% DMSO (wich helps amplifying fragments with great % of GC content). 

Example:

                                                      48 ºC            52 ºC         55 ºC        60 ºC        

1.5 mM MgCl2   ----------------------- Tube 1        Tube 2      Tube 3      Tube 4

2 mM MgCl2  ---------------------------Tube 5        Tube 6      Tube 7       Tube 8 

2 mM MgCl2 + 10% DMSO  ------ Tube 9         Tube 10     Tube 11     Tube 12

3 mM MgCl2 -------------------------- Tube 13       Tube 14      Tube 15    tube 13

Be sure to add 1 negative control (may be you could put it in the 55 ºC 2 mM MgCl2 + 10% DMSO)

If you find the amplicon you are searching in one reaction you could keep on adjusting the paremeters more tight. 

Regards

Respected sir,

re-PCR condition: Initial denaturation at 95°C for 3 minutes, denaturation at 95°C for 1 minute, annealing at 60.8°C for 30 seconds and extension at 72°C for 1 minute, and a final extension for 15 minutes with 30 cycles. Below attached picture is the DGGE gel, in that I took the single sample (marked with a black line) and did

Below attached picture is the DGGE gel, from that I excised a single band (marked with a black line) and incubated with 30ul nuclease free water overnight at 4°C. After that, I did PCR by taking various concentrations of samples like 1ul, 2ul, 3ul, 5ul. Now I couldn't get a single amplicon. Whatever you suggest let me try in the forthcoming days. 

 Mr. Martin, I extracted the samples by phenol: chloroform method and whatever you said, I will do once.

 My sincere gratitude to  both of you for giving valuable suggestions.

Sorry Santhana  I misunderstood and thought you were running a checker gel after dgge had given you an interesting result and did not realise you were extracting from stained PAG. I suspect that what may be happening here is that the amplified dna in the gel has become partially single stranded ( that is why it stopped moving in the gel ) and extraction has broken the single strand so there is very little complete dna with primer sequence at both ends . Possibly if you start with more material eg mash up the acrylamide then centrifuge and try pcr a very small volume  or phenol extract and precipitate with a carrier like glycogen or rna to bulk up the precipitate to help it to come out of solution as you do not have very much then try pcr it might work but you could also try re amplifying from the wet tube that you did the first amplification in....just add water even if there is nothing visible and re amplify or now that you know which sample is interesting can you go back to the original sample and amplify with the non clamped primers ten sequence if that is what you want to do

Respected sir,

Thank you for your suggestion and for your kind notice that, today I have done Nano dropping to quantify the amount of  DNA present in the PAG extract. I got

if that figure is correct you have a lot of material...this is quite a short pcr product so there are many molecules in 10ng but nanodrop readings can be inaccurate at low values and it needs accurate zeroing. good luck

paul

Respected sir,

             Thank you so much for spending your valuable time for solving the issue. 

Sambrook is having method to purify bands from PAGE gels you can follow that, usually after extraction concentration will be very low. Hence to increase the concentration you can load more sample to get more intense band in gel or you accumilate such expected gel bands by multiple loading then proceed for purification from gel. But elute the product in  less amount of elution buffer. Rest pcr conditions will be same. In first round of amplification amplicon intensity may be light but you can reply pcr amplified proudct again by using it as template. good luck