LPA Carrier for precipitation of small amounts of nucleic acids

The addition of linear polyacrylamide (LPA) to a nucleic acid precipitation can enhance recovery of small amounts of DNA or RNA*. LPA is an inert carrier, and is reported to not interfere with enzymatic activities, unlike some other popular carriers such as tRNA.

To an ethanol (or propanol) precipitation, add 10-20 ug of LPA

To produce a 5 ug/ul stock of linear polyacrylamide (from T. Fitzwater, ref 1, below):

In a 50 mL Falcon 2098 centrifuge tube, mix in the order listed (makes a lot, so may want to scale down to 10%):

    1. 0.25 g acrylamide (no bis-acrylamide)
    2. 4.25 mL of Millipure water
    3. 200 µL of 1 M Tris-HCl, pH 8.0
    4. 33 µL of 3 M sodium acetate, pH 7.5
    5. 10 µL of 0.5 M EDTA
    6. 50 µL of 10% APS (prepare 10% solution in sterile di-water)
    7. 5 µL of TEMED and mix well to begin polymerization; leave at room temperature 30 mins.

Add 12.5 mls 95% ETOH to precipitate approximately 5 mins.

Drain off liquid from precipitated LPA. Remove as much liquid as possible with pipet (squash pellet).

Wash pellet with 70% ETOH, then remove as much liquid as possible with pipet (squash pellet).

Air dry about 10 mins.

Add sterile di-water to LPA, let resuspend overnight with gentle shaking. Make final volume 50 mls. Concentration of LPA is 250 mg/50 mls, or 5 mg/ml, or 5 ug/ul.

Aliquot and store at -20.

Use 3 ul in a large eppy for ethanol precipitations (15 ug LPA). Be carful to not lose pellet, as it may not stick to tube as tightly as with nucleic acid carriers.

Another web protocol:

1. See LPA recipe and notes from tfitzwater (Tim Fitwater):

2. http://ubik.microbiol.washington.edu/protocols/arrays/linearacrylamide.htm

*Gaillard, C. and Strauss, F. (1990) Nucleic Acids Res. 18, 378