What are attB and attP sites?
The attP site consists of a 234-bp DNA fragment containing five scattered putative Int-binding sites (4) whereas the attB site requires only a 16-bp sequence to be functional. The mv4 minimal attB site is the shortest attB sequence described to date.
What is tyrosine integrase?
The integrases are a diverse family of tyrosine recombinases which rearrange DNA duplexes by means of conservative site-specific recombination reactions.
What are the two families of conservative site-specific Recombinases?
Only two classes of conservative site-specific recombinases have been discovered so far: the invertase/resolvase family (also called the serine family) and the lambda integrase family (also called the tyrosine family).
How do recombinases work?
Recombinases are enzymes that catalyse site-specific recombination events within DNA; for example, genetic recombination during meiosis in which recombination serves to generate new combinations of alleles on chromosomes.
What is a attP site?
The attP site (5′-GTGCCCCAACTGGGGTAACCTttGAGTTCTCTCAGTTGGGGG-3′) is one of the two target sites of the phiC31:int integrase. phiC31:int mediates recombination between two non-identical target sites, attP and attB, each of which contains a short integration core flanked by imperfect inverted repeats.
What is attL and attR?
The attL and attR sites are substrates for the excision reaction, and each is composed of one B-type half-site and one P-type half-site.
What is a phage integrase?
Phage integrases are enzymes that mediate unidirectional site-specific recombination between two DNA recognition sequences, the phage attachment site, attP, and the bacterial attachment site, attB.
What does a tyrosine recombinase do?
Tyrosine site-specific recombinases (T-SSRs) are DNA modifying enzymes that bind, cleave, strand exchange, and rejoin DNA at their respective, typically palindromic, recognition target sites.
Which enzyme plays an role in site-specific recombination?
The insertion of a circular bacteriophage lambda DNA chromosome into the bacterial chromosome. In this example of site-specific recombination, the lambda integrase enzyme binds to a specific “attachment site” DNA sequence on each chromosome, (more…)
What are the two proteins which create site-specific recombination?
There are two main protein families that mediate site-specific recombination in bacteria: the integrase family and the resolvase/invertase family [68].
What is the role of RAD51?
The RAD51 gene provides instructions for making a protein that is essential for repairing damaged DNA. Breaks in DNA can be caused by natural and medical radiation or other environmental exposures, and also occur when chromosomes exchange genetic material in preparation for cell division.
What are loxP sites?
The loxP site is a 34 bp sequences consisting of a two 13 bp inverted and palindromic repeats and 8 bp core sequences (Figure 1A). Figure 1. Mechanism of Cre-loxP system. (A) An overview of Cre-loxP system. 38 kDa Cre recombinase recognizes the loxP sites of specific 34 bp DNA sequences.
How does Gateway cloning work?
The Gateway cloning system uses two different enzyme mixtures, each of which performs a different type of recombination reaction. The BP clonase enzyme mix recombines attB sites with attP sites, generating attL and attR sites; whereas the LR clonase enzyme mix catalyzes the reverse reaction (Fig.
What are Gateway vectors?
Gateway vectors contain modified versions of the att sites so that scientists can easily clone in their desired DNA sequences. Gateway technology relies on the two reactions described below: The BP Reaction takes place between the attB sites flanking the insert and the attP sites of the donor vector.
What is an attP site?
What is an ATT site?
The minimal phage att site is composed of approximately 240-base pairs and four distinct binding sites for Int protein, at least three of which are crucial for function. This ‘donor site’ recombines efficiently with a smaller ‘recipient site’ that lacks the extensive interactions with Int protein.
What is recombination system?
A recombination system has been developed for efficient chromosome engineering in Escherichia coli by using electroporated linear DNA. A defective λ prophage supplies functions that protect and recombine an electroporated linear DNA substrate in the bacterial cell.
Is Cre a tyrosine recombinase?
Cre recombinase is a tyrosine recombinase enzyme derived from the P1 bacteriophage. The enzyme uses a topoisomerase I-like mechanism to carry out site specific recombination events.
What components are needed for site-specific recombination?
In its strictest definition (as used in this review), site-specific recombination involves (a) two DNA partners, (b) a specialized recombinase protein that is responsible for recognizing the sites and for breaking and rejoining the DNA, and (c) a mechanism that involves DNA breakage and reunion with conservation of the …
Why is site-specific recombination important?
Site-specific recombination systems mediate certain types of genetic exchange in prokaryotes for processes that are essential for growth and adaptation mechanisms such as control of gene expression.
What are the types of recombination?
At least four types of naturally occurring recombination have been identified in living organisms: (1) General or homologous recombination, (2) Illegitimate or nonhomologous recombination, (3) Site-specific recombination, and (4) replicative recombination.
What protein does RAD51 encode?
DNA repair protein RAD51 homolog 1 is a protein encoded by the gene RAD51. The enzyme encoded by this gene is a member of the RAD51 protein family which assists in repair of DNA double strand breaks. RAD51 family members are homologous to the bacterial RecA, Archaeal RadA and yeast Rad51.
What type of gene is RAD51?
Protein Coding gene
RAD51 (RAD51 Recombinase) is a Protein Coding gene. Diseases associated with RAD51 include Fanconi Anemia, Complementation Group R and Mirror Movements 2. Among its related pathways are Endometrial cancer and Cell Cycle, Mitotic.
What is loxP used for?
General principle of Cre-loxP system
Cre-loxP system is a widely used powerful technology for mammalian gene editing. This system has advantages which is very simple manipulation and do not require additional factors for efficient recombination [1].
Where do you put a loxP site?
Generally, two loxP sites are inserted into the introns flanking an essential exon of the target gene.