Human Genome Project sequencing
Making libraries
The first step is to make a library of the DNA. Like your local library, a DNA library is a collection of documents: in this case, they are DNA molecules. In the Human Genome Project, the DNA library made from pieces of human DNA joined to special DNA sequences to produce Bacterial Artificial Chromosomes or BACs, each of which can contain about 200,000 base pairs of DNA.
The BACs are then positioned on the maps produced earlier, so researchers know which part of the genome is in which BAC: they then begin sequencing them. Each BAC is cut into smaller fragments, each about 2000 base pairs long. By breaking lots of copies of the BAC at random, a large set of overlapping smaller fragments is produced.
The smaller fragments are then joined to short DNA sequences that allow them to be grown in bacteria in the lab. The new, circular pieces of DNA are called plasmids or pUC clones. The areas where the two pieces of DNA meet - the DNA to be sequenced and the known DNA - are the sections sequenced.
After growing the bacteria, the DNA is isolated and the DNA sequencing reactions are carried out. DNA sequencing reactions can only 'read' about 500 bases at a time, so lots of reads are needed to cover each 200,000-base pair BAC.
