Help!What's wrong with the endonuclease digestion of total DNA of - (Mar/08/2005 )

Total DNA from activated sludge was extracted and purified to amplify the fragment of 16SrRNA by using eubacterial universal primers. Some of PCR products were directly using the HaeIII to produce RFLP patterns.Unfortunately,by trying different systems and ways of endonuclease digestion,I can not obtain any clear and distinct DNA patterns, What I get is faint patterns
and are almost invisible. The digestion system is PCR product 8ul,endonuclease HaeIII 2U,PCR buffer 1ul.However, using this system in bacterial isolates,I can get a clear pattern.What's wrong with my endonuclease digestion, I attemp different reaction system and electrophoresis conditions,what I get is the faint,unclear DNA pattern,which I post in attachment.
Could anybody can do me a favor and tell me what's the matter and how to solve this problem! Thanks a lot, I am so eager to your solution.

I do not know what the composition of the PCR buffer being used is. However I am willing to bet that if it were checked against the NEB buffer, the PCR buffer would most likely resemble a high salt buffer. Not to mention that most PCR buffers have a pH around 8 to 9. While NEB buffers are around pH7.5

Thus I have two suggestion,

1- Percipitate the PCR product with aid of dextran, then resuspend in water. Do add the buffer recommended by NEB, as microcharm has stated. Add the enzyme last and digest. There is a risk that the PCR product is too dilute to percipitate efficiently.And time consuming too for multiple samples

2-Alternative, use the PCR DNA directly but dilute it with a larger volume of enzyme buffer solution

example
10ul PCR DNA, in PCR mix
3ul Buffer 2
0.5ul 5U HaeIII
27ul ds water

Digest for the desired time.

I would give it 2hrs and not be bothered with DNA concentration. One less thing to worry about


HaeIII
Activity in NEBuffers:
NEBuffer 1: 50%
NEBuffer 2: 100%
NEBuffer 3: 25%
NEBuffer 4: 75%