Find Overlap By Computer

Contig #1 Gap Contig #2

The first draft of the human genome appeared in early 2001.

was to sequence the DNA making up the entire human genome. When it first began in 1990, the official Human Genome Project was to be completed by 2005. In 1990, it cost $10 to determine each base of sequence, but technological advances reduced the cost of sequencing to 50 cents per base by the late 1990s. Therefore, the Human Genome Project was to cost roughly 1.5 billion dollars.

However, in 1998 an upstart private venture, Celera Genomics, led by Craig Venter, claimed that it would finish the job by the end of 2001 at a cost of a mere $200,000. To prove it was serious, Celera Genomics sequenced the entire genome of the fruit fly, Drosophila, between May and December of 1999. Three million random sequence reads were assembled to give the 180 Mb genome, demonstrating the feasibility of the shotgun approach. The private and public Human Genome Projects jointly announced that the first draft of the human genome was complete in June 2000. In fact, the Celera sequence was 99% complete whereas the public project was only 85% done. A working draft was published in February of 2001.

The official Human Genome Project proceeded by cloning large fragments of human DNA, mostly in yeast artificial chromosomes (YACs) and bacterial artificial chromosomes (BACs), and mapping them to their chromosomal locations. Only then were the large mapped fragments broken up for shotgun sequencing. Although this makes assembly easier, cloning consumes time and money. Instead, Celera used the

FIGURE 24.21 Closing Gaps between Contigs

To identify gaps between contigs, probes or primers are made that correspond to the ends of the contigs (pink). In (A) a new library of clones (green) is screened with end-of-contig probes. Clones that hybridize to probes from two sides of a gap are isolated. In this example a probe for the end of contig #3 (3b) and the beginning of contig #4 (4a) hybridize to the fragment shown. Therefore, the sequence of this clone should close the gap between contig #3 and #4. The second approach uses PCR (B). PCR primers that correspond to the ends of contigs are combined in random pairs and used to amplify genomic DNA. If the primer pair is within a few kilobases of each other, a PCR product is made and can be sequenced.

Contig #1 Contig #2

Contig #1 Contig #2

Make probes or primers that

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