Fractionation of DNA from marine invertebrate ( Maja Crispata, Mytilus Galloprovincialis) haemolymph by alkaline elution

Abstract

1. 1. An alkaline elution procedure for the detection of DNA damage in marine invertebrate haemolymph has been developed. 2. 2. Provided that three criteria are optimized, such as buffer composition, small filter pores (0.22 μm GVWP 025 00, Millipore), and optimal amounts of haemolymph applied, flow rates may be changed within the range of 0.2 ml/min to 0.05 ml/min without advere back-pressure on the filter and without blocking filter pores. 3. 3. Under optimal conditions, 70% of mussel haemolymph DNA, and 80% of crab haemolymph DNA will be retained on the filter 6 hr of elution, indicating shorter DNA in mussel haemolymph. 4. 4. The technique is applicable for testing the vivo effects of different compounds on DNA in marine invertebrates, and to measurements of DNA damage in naturally exposed mussels. 5. 5. This argues an important case for the use of alkaline elution technique for assessement of environmental genotoxicity, and especially for investigation of DNA damage in different marine organisms which cover a broad range in their DNA molecular weights.