Yeast Artificial Chromosome (YAC)
Yeast artificial chromosome (YAC)
The yeast artificial chromosome, which is often shortened to YAC, is an artificially constructed system that can undergo replication. The design of a YAC allows extremely large segments of genetic material to be inserted. Subsequent rounds of replication produce many copies of the inserted sequence, in a genetic procedure known as cloning .
The reason the cloning vector is called a yeast artificial chromosome has to do with the structure of the vector. The YAC is constructed using specific regions of the yeast chromosome. Yeast cells contain a number of chromosomes ; organized collections of deoxyribonucleic acid (DNA ). For example, the yeast Saccharomyces cerevisae contains 16 chromosomes that contain varying amounts of DNA. Each chromosome consists of two arms of DNA that are linked by a region known as the centromere. As the DNA in each arm is duplicated, the centromere provides a region of common linkage. This common area is the region to which components of the replication machinery of the cell attach and pull apart the chromosomes during the cell division process. Another region of importance is called the telomere. The end of each chromosome arm contains a region of DNA called the telomere. The telomere DNA does not code for any product, but serves as a border to define the size of the chromosome. Finally, each chromosome contains a region known as the origin of replication. The origin is where a molecule called DNA polymerase binds and begins to produce a copy of each strand of DNA in the double helix that makes up the chromosome.
The YAC was devised and first reported in 1987 by David Burke, who then also reported the potential to use the construct as a cloning vehicle for large pieces of DNA. Almost immediately, YACs were used in large-scale determination of genetic sequences, most prominently the Human Genome Project.
YAC contains the telomere, centromere, and origin of replication elements. If these elements are spliced into DNA in the proper location and orientation, then a yeast cell will replicate the artificial chromosome along with the other, natural chromosomes. The target DNA is flanked by the telomere regions that mark the ends of the chromosome, and is interspersed with the centromere region that is vital for replication. Finally, the start site for the copying process is present. In essence, the yeast is fooled into accepted genetic material that mimics a chromosome.
The origin of the DNA that is incorporated into a YAC is varied. DNA from prokaryotic organisms such as bacterial or from eukaryotes such a humans can be successfully used. The power of YACs is best explained by the size of the DNA that can be copied. Bacteria are also capable of cloning DNA from diverse sources, but the length of DNA that a bacterium can handle is up to 20 times less than that capable of being cloned using a YAC.
The engineered YAC is put back into a yeast cell by chemical means that encourage the cell to take up the genetic material. As the yeast cell undergoes rounds of growth and division, the artificial chromosome is replicated as if it were a natural chromosomal constituent of the cell. The result is a colony of many genetically identical yeast cells, each containing a copy of the target DNA. The target DNA has thus been amplified in content. Through a subsequent series of procedures, DNA can then be isolated from the rest of the DNA inside the yeast cells.
Use of different regions of DNA in different YACs allows the rapid determination of the sequence, or order of the constituents, of the DNA. YACs were invaluable in this regard in the sequencing of the human genome, which was completed in preliminary form in 2001 The human genome was broken into pieces using various enzymes . Each piece could be used to construct a YAC. Then, sufficient copies of each piece of the human genome could be generated so that automatic sequencing machines would have enough material to sequence the DNA.
Commonly, the cutting enzymes are selected so that the fragments of DNA that are generated contain overlapping regions. Once the sequences of all the DNA regions are obtained the common overlapping regions allow the fragment sequences to be chemically bonded so that the proper order and the proper orientation is generated. For example, if no overlapping regions were present, then one sequence could be inserted backwards with respect to the orientation of its neighbouring sequence.
See also Chromosomes, prokaryotic; Gene amplification; Yeast genetics