Vector is a means by which the deoxyribonucleic acid fragment is carried into the cell of the host. A cloning vector is a tiny segment of DNA into which a foreign DNA portion can be inserted. Expression vectors are the vectors that include the necessary regulatory sequences particularly used for the expression of a specific gene into proteins inside the target cell.' YAC's or Yeast artificial chromosomes are shuttle-vectors, ( a shuttle vector is one that is able to replicate in other than one host organisms or 2 dissimilar cell types that is a prokaryotic and a eukaryotic cell) that have the ability to amplify in bacteria and can be utilized for the manipulation and cloning of huge DNA inserts in Saccharomyces cerevisiae (yeast). These vectors include features necessary for stability and replication in yeast cells.
Artificial chromosomes can be utilized to clone large segments of DNA in yeast cells. Initially depicted in the year 1983 by Szostak and Murray, a YAC is an artificially built chromosome and includes the centromere, telomere and autonomously replicating sequences essential for replication and conservation in yeast cells. YAC's are of quite small size (nearly 12 kb) and of spherical form when they are amplified or manipulated in bacteria, but are altered linear and of very huge size, that is quite a lot of hundreds of kilobases (kb), when introduced as cloning vectors in yeast. The later procedure involves cleavage at specifically located sites by 2 restriction enzymes, which break them in two linear DNA arms. These are consequently ligated with the suitable DNA insert before transformation into recipient yeast cells which are converted to spheroplasts (cell from which the cell wall has been almost completely removed ). Then the telomere sequences are inserted by the telomerase enzymes in the cells.
In the linear structure, these specific vectors contain all three essential structural components necessary for performing like wild yeast chromosomes.
- ARS - An autonomously replicating sequence required for replication.
- CEN - A centromere for segregation at cell division.
- TEL - Two telomeres for conservation
- In addition, their ability to admit large DNA inserts makes possible for them to attain the minimum size (150 kb) necessary for chromosome-like constancy and for stability of transmission in yeast cells.
Applications of YACs include;
- Withholding of DNA inserts thousands of kilobases in size and steady preservation of cloned eukaryotic DNA due to the compatibility with the yeast replication mechanisms.
- They are open to to large-scale plasmid amplification in bacteria and to construction of precise genetic changes within the exogenous DNA sequences by using the dedicated and well-organized yeast mechanism of homologous recombination.
- Yeast artificial chromosomes correspond to the top tools for the learning of eukaryotic genomes and for mobilisation of huge genetic elements among bacteria and eukaryotes.
- YACs function like wildiy existing chromosomes, in regard that they are of the appropriate size, revealing analogous stability.
- YACs can be manipulated straightly by traditional genetic engineering as well as by modern recombination mediated genetic engineering technology.
- Appropriately modified, YACs can be utilized in many different organisms, for cloning or genome analysis.
- Chromosomal translocation (chromosome abnormality that occurs due to rearrangement of parts among non homologous chromosomes) can be studied by means of disposable YACs that do not contain genetic information necessary for cell function.
- Artificial chromosome vectors have made easier the mapping and sequencing of complex genomes at an progressively more rapid rate. Conventional bacterial cloning agents have remained significant for the learning of rather short clones, but for the cloning of very huge DNA pieces artificial chromosomes have totally replaced prior bacterial systems, counting the lambda phage-based cosmid vectors.
Artificial chromosomes can be suitably constructed and manipulated in yeast cells using in vivo (within a living organism) homologous recombination, a new process called as recombination mediated genetic engineering (recombineering). YACs can also be utilized for engineering genetic determinants of novel biochemical pathways for manufacture of secondary metabolites and for heterologous expression of proteins. YACs can be modified with the suitable selectable markers and transferred to cells of various organisms permitting the production of transgenic animals. Ultimately, YACs are greatly engaged in the construction of fullâ€scale genomic libraries, for mapping and functional analysis.
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