A brief account of YAC, BAC, HAC
1. YAC (Yeast Artificial Chromosome):-
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> It was first described in 1983 by Murray and Szostak.
> These are artificially constructed vectors. The system can undergo replication and has the capability to insert foreign DNA sequences.
> Components such as the autonomously replication system (ARS), centromere and telomeres are taken from the yeast Saccharomyces cerevisiae for the construction of vectors.
> The desired gene sequence can be inserted into these vectors and then put back into yeast; the yeast machinery cannot differentiate between their own and foreign genes; hence the foreign gene is also replicated.
> The desired gene sequence can be inserted into these vectors and then put back into yeast; the yeast machinery cannot differentiate between their own and foreign genes; hence the foreign gene is also replicated.
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Features:-
- YAC vectors are DNA constructs that are used for cloning DNA in yeasts.
- The amount of DNA that can be cloned into a YAC is, on average, from 200 to 500 kb. However, as much as 1 Mb can be cloned into a YAC.
- They often show chimerism. It contain DNA in a single clone from different locations in the genome. It is a problem encountered in constructing and using YAC libraries is that they typically contain clones that are chimeric.
- It is linear.
- Yeast machinery has post-translational mechanisms that are useful in the expression of eukaryotic proteins.
- Only one vector occurs per yeast cell.
- Less stable.
- The efficiency of cloning is low (about 1000 clones are obtained per microgram of vector and insert DNA).
- Advantage:- Many sequences that are unstable, underrepresented, or absent when cloned into prokaryotic systems, remain stable and intact in YAC clones.
- Disadvantage:- There are chances of gene deletion and gene recombination or inversion in the inserted gene sequence.
- Applicable in gene mapping and chromosome walking.
2. BAC (Bacterial Artificial Chromosome):-
> It was first developed by Shizuya in 1992.
> These are DNA constructs that are used for transformation and cloning in bacterial cells, mostly E.coli. Functional fertility plasmids or F-plasmids are used for the vector construction.
> The gene components included are rep6 for plasmid regulation, a selectable marker for antibiotic resistance, parA and parB for F-plasmid DNA, and T7 and Sp6 for transcription of inserted genes.
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Features:-
- BAC vectors are DNA constructs that are used for cloning DNA in bacteria.
- These are simple plasmid which is designed to clone very large DNA fragments ranging in size from 75 to 300 kb.
- No chimerism seen.
- It is circular.
- Bacterial machinery does not have post-translational mechanism, hence the expression of eukaryotic proteins becomes difficult.
- 1-2 vectors are found per bacterial cell.
- More stable.
- Avantage:- The generation of BACs is quicker and more efficient, and also, it gives a better chromosome coverage map. Hence, today, BAC is preferred over YAC because they are more efficient.
- Applicable in modelling genetic diseases and human genome project.
3. HAC (Human Artificial Chromosome):-
> It is first described by Harrington (1997).
> It synthesized by combining portions of alpha satellite DNA with telomeric DNA and genomic DNA into linear micro chromosomes.
> HACs are microchromosomes that can act as a new chromosome in a population of human cells. It means instead of 46 chromosomes, the cell could have 47 chromosomes with the 47th being very small,
and able to carry new genes introduced by human researchers.
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> HACs range in size from 6 to 10 Mb that carry new genes introduced by human researchers.
> HACs can be used as vectors in transfer of new genes, studying their expression and mammalian chromosomal function can also be elucidated using these microchrosomes in mammalian system.
> Human artificial chromosomes are extrachromosomal DNA fragments that act as a new chromosome within the human cell.
> The use of human artificial chromosomes has increased with advances in genetic engineering as it helps overcome problems commonly associated with traditional vector systems.
> HACs can exist as single copy episomes without integration into the host chromosomes allowing long-term stable maintenance.
> Besides, there is no upper limit in the size of the DNA insert to be incorporated into a HAC as entire genomic units can be used to mimic the natural gene expression.
> In spite of numerous advantages, HACs have only been used for studies related to the structure and function of human kinetochores.
> Limitations associated with HACs are due to technical difficulties during gene loading and ill-defined structures of the vectors.