Bacterial artificial chromosome
Encyclopedia
A bacterial artificial chromosome (BAC) is a DNA construct
, based on a functional fertility plasmid
(or F-plasmid), used for transforming
and cloning
in bacteria
, usually E. coli. F-plasmids play a crucial role because they contain partition genes that promote the even distribution of plasmids after bacterial cell division. The bacterial artificial chromosome's usual insert size is 150-350 kbp
, but can be greater than 700 kbp. A similar cloning vector
called a PAC
has also been produced from the bacterial P1-plasmid.
BACs are often used to sequence
the genome of organisms in genome project
s, for example the Human Genome Project
. A short piece of the organism's DNA
is amplified as an insert in BACs, and then sequenced. Finally, the sequenced parts are rearranged in silico
, resulting in the genomic sequence of the organism.
parA and parB: for partitioning F plasmid DNA to daughter cells during division and ensures stable maintenance of the BAC.
A selectable marker: for antibiotic resistance; some BACs also have lacZ at the cloning site for blue/white selection
.
T7 & Sp6: phage promoters for transcription of inserted genes.
associated with Down syndrome. There have also been instances when they have been used to study specific oncogene
s associated with cancers. They are transferred over to these genetic disease models by electroporation/transformation, transfection with a suitable virus or microinjection. BACs can also be utilised to detect genes or large sequences of interest and then used to map them onto the human chromosome using BAC arrays
. BACs are preferred for these kind of genetic studies because they accommodate much larger sequences without the risk of rearrangement, and are therefore more stable than other types of cloning vectors.
, poxviruses
and coronaviruses
more accessible. Molecular studies of these viruses can now be achieved using genetic approaches to mutate the BAC while it resides in bacteria. Such genetic approaches rely on either linear or circular targeting vectors to carry out homologous recombination
.
DNA construct
A DNA construct is an artificially constructed segment of nucleic acid that is going to be "transplanted" into a target tissue or cell...
, based on a functional fertility plasmid
Plasmid
In microbiology and genetics, a plasmid is a DNA molecule that is separate from, and can replicate independently of, the chromosomal DNA. They are double-stranded and, in many cases, circular...
(or F-plasmid), used for transforming
Transformation (genetics)
In molecular biology transformation is the genetic alteration of a cell resulting from the direct uptake, incorporation and expression of exogenous genetic material from its surroundings and taken up through the cell membrane. Transformation occurs naturally in some species of bacteria, but it can...
and cloning
Cloning
Cloning in biology is the process of producing similar populations of genetically identical individuals that occurs in nature when organisms such as bacteria, insects or plants reproduce asexually. Cloning in biotechnology refers to processes used to create copies of DNA fragments , cells , or...
in bacteria
Bacteria
Bacteria are a large domain of prokaryotic microorganisms. Typically a few micrometres in length, bacteria have a wide range of shapes, ranging from spheres to rods and spirals...
, usually E. coli. F-plasmids play a crucial role because they contain partition genes that promote the even distribution of plasmids after bacterial cell division. The bacterial artificial chromosome's usual insert size is 150-350 kbp
Base pair
In molecular biology and genetics, the linking between two nitrogenous bases on opposite complementary DNA or certain types of RNA strands that are connected via hydrogen bonds is called a base pair...
, but can be greater than 700 kbp. A similar cloning vector
Cloning vector
A cloning vector is a small piece of DNA into which a foreign DNA fragment can be inserted. The insertion of the fragment into the cloning vector is carried out by treating the vehicle and the foreign DNA with a restriction enzyme that creates the same overhang, then ligating the fragments...
called a PAC
P1-derived artificial chromosome
The P1-derived artificial chromosome are DNA constructs that are derived from the DNA of P1 bacteriophage. They can carry large amounts of other sequences for a variety of bioengineering purposes...
has also been produced from the bacterial P1-plasmid.
BACs are often used to sequence
Sequencing
In genetics and biochemistry, sequencing means to determine the primary structure of an unbranched biopolymer...
the genome of organisms in genome project
Genome project
Genome projects are scientific endeavours that ultimately aim to determine the complete genome sequence of an organism and to annotate protein-coding genes and other important genome-encoded features...
s, for example the Human Genome Project
Human Genome Project
The Human Genome Project is an international scientific research project with a primary goal of determining the sequence of chemical base pairs which make up DNA, and of identifying and mapping the approximately 20,000–25,000 genes of the human genome from both a physical and functional...
. A short piece of the organism's DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
is amplified as an insert in BACs, and then sequenced. Finally, the sequenced parts are rearranged in silico
In silico
In silico is an expression used to mean "performed on computer or via computer simulation." The phrase was coined in 1989 as an analogy to the Latin phrases in vivo and in vitro which are commonly used in biology and refer to experiments done in living organisms and outside of living organisms,...
, resulting in the genomic sequence of the organism.
Common gene components
oriS, repE - F: for plasmid replication and regulation of copy number.parA and parB: for partitioning F plasmid DNA to daughter cells during division and ensures stable maintenance of the BAC.
A selectable marker: for antibiotic resistance; some BACs also have lacZ at the cloning site for blue/white selection
Blue white screen
The blue-white screen is a screening technique that allows for the detection of successful ligations in vector-based gene cloning. DNA of interest is ligated into a vector. The vector is then transformed into competent cell . The competent cells are grown in the presence of X-gal...
.
T7 & Sp6: phage promoters for transcription of inserted genes.
Inherited disease
BACs are now being utilized to a greater extent in modelling genetic diseases, often alongside transgenic mice. BACs have been useful in this field as complex genes may have several regulatory sequences upstream of the encoding sequence, including various promoter sequences that will govern a gene's expression level. BACs have been used to some degree of success with mice when studying neurological diseases such as Alzheimer's disease or as in the case of aneuploidyAneuploidy
Aneuploidy is an abnormal number of chromosomes, and is a type of chromosome abnormality. An extra or missing chromosome is a common cause of genetic disorders . Some cancer cells also have abnormal numbers of chromosomes. Aneuploidy occurs during cell division when the chromosomes do not separate...
associated with Down syndrome. There have also been instances when they have been used to study specific oncogene
Oncogene
An oncogene is a gene that has the potential to cause cancer. In tumor cells, they are often mutated or expressed at high levels.An oncogene is a gene found in the chromosomes of tumor cells whose activation is associated with the initial and continuing conversion of normal cells into cancer...
s associated with cancers. They are transferred over to these genetic disease models by electroporation/transformation, transfection with a suitable virus or microinjection. BACs can also be utilised to detect genes or large sequences of interest and then used to map them onto the human chromosome using BAC arrays
DNA microarray
A DNA microarray is a collection of microscopic DNA spots attached to a solid surface. Scientists use DNA microarrays to measure the expression levels of large numbers of genes simultaneously or to genotype multiple regions of a genome...
. BACs are preferred for these kind of genetic studies because they accommodate much larger sequences without the risk of rearrangement, and are therefore more stable than other types of cloning vectors.
Infectious disease
The genomes of several large DNA viruses and RNA viruses have been cloned as BACs. These constructs are referred to as "infectious clones", as transfection of the BAC construct into host cells is sufficient to initiate viral infection. The infectious property of these BACs has made the study of many viruses such as the herpesvirusesHerpesviridae
The Herpesviridae are a large family of DNA viruses that cause diseases in animals, including humans. The members of this family are also known as herpesviruses. The family name is derived from the Greek word herpein , referring to the latent, recurring infections typical of this group of viruses...
, poxviruses
Poxviridae
Poxviruses are viruses that can, as a family, infect both vertebrate and invertebrate animals.Four genera of poxviruses may infect humans: orthopox, parapox, yatapox, molluscipox....
and coronaviruses
Coronaviridae
Coronaviruses are enveloped, single-stranded, positive-sense RNA viruses with club-shaped surface about 120-160 nm in diameter that resemble a “corona”.-Virology:...
more accessible. Molecular studies of these viruses can now be achieved using genetic approaches to mutate the BAC while it resides in bacteria. Such genetic approaches rely on either linear or circular targeting vectors to carry out homologous recombination
Homologous recombination
Homologous recombination is a type of genetic recombination in which nucleotide sequences are exchanged between two similar or identical molecules of DNA. It is most widely used by cells to accurately repair harmful breaks that occur on both strands of DNA, known as double-strand breaks...
.
See also
- Human artificial chromosomeHuman artificial chromosomeA human artificial chromosome is a microchromosome that can act as a new chromosome in a population of human cells. That is, instead of 46 chromosomes, the cell could have 47 with the 47th being very small, roughly 6-10 megabases in size, and able to carry new genes introduced by human researchers...
- Yeast artificial chromosomeYeast artificial chromosomeA yeast artificial chromosome is a vector used to clone DNA fragments larger than 100 kb and up to 3000 kb. YACs are useful for the physical mapping of complex genomes and for the cloning of large genes...
- FosmidFosmidFosmids are similar to cosmids but are based on the bacterial F-plasmid. The cloning vector is limited, as a host can only contain one fosmid molecule. Fosmids are 40 kb of random genomic DNA...
- CosmidCosmidA cosmid, first described by Collins and Hohn in 1978, is a type of hybrid plasmid that contains cos sequences, DNA sequences originally from the Lambda phage. Cosmids can be used to build genomic libraries....
External links
- The Big Bad BAC: Bacterial Artificial Chromosomes — a review from the Science Creative Quarterly
- Empire Genomics (company that sells BAC clones from genomic libraries)
- Amplicon Express (company that makes custom BAC libraries)