Home Molecular Biology Extraction Of High Molecular Weight DNA From Microbial Mats l Protocol Preview
Steps
  1. 1 Homogenize microbial mat sample 01:41
  2. 2 Extract microbial cells from mat 00:12
  3. 3 Lyse cells using freeze-thaw and chemicals 00:21
  4. 4 Remove cell debris and RNA 00:29
  5. 5 Extract and concentrate DNA 00:39
  6. 6 Precipitate and purify DNA 00:48
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Extraction Of High Molecular Weight DNA From Microbial Mats l Protocol Preview

Protocol
Difficulty
intermediate

Steps

1
Homogenize microbial mat sample

Mix approximately 30 grams of microbial mat thoroughly using a sterile grinding pestle without grinding until no lumps remain visible in the mixture.

▶ 01:41
2
Extract microbial cells from mat

Separate microbial cells from the homogenized mat background matrix using the prepared sample.

▶ 00:12
3
Lyse cells using freeze-thaw and chemicals

Apply freeze-thaw cycles combined with chemical lysis methodology to break open the microbial cells and release DNA.

▶ 00:21
4
Remove cell debris and RNA

Perform organic extractions and RNase treatment to eliminate cell debris and RNA from the lysate, leaving DNA in solution.

▶ 00:29
5
Extract and concentrate DNA

Isolate DNA from the treated solution using extraction methodology to separate it from other contaminants.

▶ 00:39
6
Precipitate and purify DNA

Perform DNA precipitation and purification steps to obtain high molecular weight DNA of optimal quality and concentration.

▶ 00:48

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critical
DNA Fragmentation and Size Reduction
Extracted DNA shows reduced fragment size on gel electrophoresis or pulse-field gel analysis. UHMW DNA appears degraded with smaller molecular weight distribution than expected.
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