"My primers amplification got many time primer dimer, and i did not get my desired band in pcr amplification."

Perhaps it is time to design new primers or try a different PCR profile such as 'touchdown' PCR, nested PCR or use the same annealing/elongation temps in the 68°C-72°C range.

"I want to cut the genomic DNA from the gel, and use it as the template for a new PCR reaction."

Not sure why you would want to do this as gDNA would simply be a smear on your gel from top to bottom. Also, re-isolating gDNA from a gel would probably not fix problem #1. If it is actually the PCR band that you want to gel isolate, then fix problem #1. If you want better gDNA template because you suspect impurities for the above PCR, there are better ways to clean it up prior to your PCR, e.g. phenol/chloroform extraction or alcohol precipitation, that doesn't require gel isolation.

Take excised agarose slice

Place inside a 0.5ml eppendorf tube which has been deliberately been pierced with a needle

So sometimes the agarose slice can be squeezed through a wide bore needle using a syringe into the inner tube or recovery facilitated by adding glass wool to this inner tube ( this was done routinely in the early 90s before the advent of purification kits):

http://www.jcancer.org/v03p0093.htm

place slice in pierced 0.5ml tube into a 1.5ml eppendorf

Spin this two tube arrangement at 13,000rpm for 10 minutes during which time your aqueous component plus DNA will make its way via the hole into the 1.5ml eppendorf

Analogous with the above the eluate can often be used successfully in secondary PCR. Occasionally however this is not true by virtue of excessive salt and residual mucoploydacharide so desalt the eluate by phenol chloroform extraction and ethanol ppt

http://www.oardc.ohio-state.edu/stockingerlab/Protocols/PhenolExtraction.pdf

Specifically

Add an equal volume of Tris buffered phenol to chloroform Isoamy alcohol (24:1)

Vortex for 1 minute

Spin for 5 min in a microfuge at 13K

remove the upper Aqueous phase gently from the top down taking care not to remove the milky interphase ( in your case mucopolysaccharide)

Transfer this upper aqueous phase to a fresh tube with chloroform iso amyl alcohol (24 parts chloroform 1 part IAA)

Vortex and repeat steps 2-4

With upper Aqueous phase add:

3 volumes of 95%-100% molecular ethanol to your sample

1/10 volume 3M sodium acetate

If amplicon size is < 100bp 1/10 1mM MgCl(2) as well

Moteover for improved recovery also add 1ul of 1mg/ml glycogen

Incubate -20C for 15 min

Spin at 13k for 15 min in a microfuge

remove most of the supernatant from the top down

Add back 1ml of ROOM TEMP 70% ethanol

Vortex

Spin 5 min to pellet insoluble material

Repeat 8. and 9.

Finally spin for 5 minutes remove most of the ethanol with a p200 down to the pellet

Spin for a further min to bring down residual ethanol

remive residue with p10 tip against the pellet

Air dry for 15 min at room temp

Resuosend touch dry pellet in 50ul of molecular grade water

Use 1ul in your PCR

Most of the time as indicated eluate derived directly from your gel slice will work in PCR but sometimes this additional purification will improve PCR efficiency

Alternatively take the the eluate spun from gel slice

Mix with an equal volume of 70% ethanol

Apply to a commercial silica DNA purification column and follow standard protocol

However leave the ethanol based wash buffer 1 minute before spinning through and repeat this ethanol based desalt wash

my only other modification is to add 30ul of water preheated to 50C to dry column; wait 5 min; spin for 1 min to elute the DNA

Repeat 5 but with 20ul of water

Pool fractions 5. and 6. and use 1ul of resulting 50ul eluate in PCR

Or simply cut the gel with desired band and add PCR water smash the agarose with a yellow tip, centrifuge it at maximum speed. Collect the liquid (DNA) above gel and use it for second round of PCR. Sometimes it works as well.

"My primers amplification got many time primer dimer, and i did not get my desired band in pcr amplification."

Perhaps it is time to design new primers or try a different PCR profile such as 'touchdown' PCR, nested PCR or use the same annealing/elongation temps in the 68°C-72°C range.

"I want to cut the genomic DNA from the gel, and use it as the template for a new PCR reaction."

Not sure why you would want to do this as gDNA would simply be a smear on your gel from top to bottom. Also, re-isolating gDNA from a gel would probably not fix problem #1. If it is actually the PCR band that you want to gel isolate, then fix problem #1. If you want better gDNA template because you suspect impurities for the above PCR, there are better ways to clean it up prior to your PCR, e.g. phenol/chloroform extraction or alcohol precipitation, that doesn't require gel isolation.

I don't know if you can recover genomic DNA from an agarose gel. I wouldn't bother to try unless it's an irreplaceable sample. Just try your PCR again with your saved genomic DNA (I'm assuming you didn't use the entire genomic DNA sample in one PCR reaction).

Hi.

I guess your problem, as someone else already told here, is associated with a not completely standardized PCR. Primer dimers occur quite often but do not disturb the PCR result.

If you're always getting not specific bands, it is standardization requirements which you did not get that are doing it, or bad primer design. In both cases, gel extracted DNA (with all different salts or re-purified) doesn't seem to be what is going to solve your problem.

If I understood it not the right way ... then:

I guess Saima Shakil Malik already gave you lots of nice ideas to try. One other easy way that sometimes works fine is to put the gel slice into a clean alluminium foil, some PCR-grade water and freeze it. After, put it over a microfuge tube and let the water come to the new tube. Sometimes works fine ... Can be beneficial to desalt and re-purify for even better results.

Hope it helps.

I agree, it seems that yur problem could be found within your protocol/procedure or may be due to the experimentator.