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New Protocols for Isolating High- Molecular-Weight Genomic DNA

ith higher packaging efficiencies, make the RecoverEase DNA isolation kit the recommended method for purifying high-molecular-weight genomic DNA from blood, bone marrow, and cultured cells.

Big Blue Transgenic Rodent Mutation Assay and Cell Culture System

Genomic DNA isolated with this kit is ideal for recovering lambda shuttle vectors in transgenic in vivo mutation assay systems, such as Stratagenes Big Blue mutation assay systems.*,3,4,5 An important step in these mutation assays is retrieving large quantities of shuttle vector from the animals genomic DNA for subsequent analysis of mutations within a target region of the vector. The shuttle vector in Big Blue was derived from bacteriophage lambda and can be retrieved from the animal by subjecting the genomic DNA to the specialized in vitro lambda Transpack packaging extract.**,4 Because the shuttle vector is nearly 50-kb long and is present in multiple copies within the genome of the animal, efficient packaging is contingent on genomic DNA preparations of very high-molecular weight.6 The DNA must also have sufficient purity so as to be free of substances that inhibit the packaging reaction. For these reasons, measuring packaging efficiency in DNA preparations made from shuttle vector transgenics is a good indicator of DNA size and purity. DNA yielded from using the kit averaged a 74% greater recovery of plaque-forming units (pfu) per Transpack packaging reaction over organic extracted DNA overall for liver, kidney, spleen, testes, lung, and brain in Big Blue transgenic mice.6

RecoverEase DNA Isolation Kit vs. Organic Extraction

The RecoverEase method (see Methods) was compared with the organic extraction technique in relation to DNA yield, quality, and packaging efficiency. The latter technique was taken from Stratagenes Big Blue DNA isolation kit, whic
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