In steps towards the major goal of sequencing the human genome, several investigators are developing partial maps of chromosomal DNA. Libraries consisting of clones of overlapping DNA fragments exist in several laboratories. The goal is to reconstruct the order to the fragments as they appear in the original DNA, thus obtaining sequences of overlapping fragments that cover large segments of the DNA. The overlapping fragments may be originally obtained by partial digestion of the DNA by a single enzyme or total digestion by two or more enzymes used individually. A critical requirement is that the fragments be of a size appropriate to cloning in some bacterial system. The second step is to identify as many pairs of overlapping fragments as possible; theoretically, if the library covers the chromosomal DNA and all intersecting pairs can be identified, a single sequence of overlapping fragment reaching from one end of the DNA to the other would be obtained. Identification of overlapping pairs may be made by observing a common pattern of restriction fragment lengths when each fragment of the pair is digested with a certain digest. Alternatively, the same subset of a panel of probes (complementary DNA fragments) may bind specified parts of each fragment of the pair. Using a randomly generated sequence as a model DNA, we are evaluating several of these procedures to determine which of them should lead to the highest percentage coverage of the original chromosome, with the greatest accuracy of ordering, and with the fewest number of experimental steps.
