What is actually pUC19?
pUC19 is a plasmid cloning vector which was first discovered by Joachim Messing and his team. It is a widely used vector for recombination, or for introducing foreign DNA into cell. The word “pUC” is derived from “p” denoting the word plasmid and the abbreviation for the University Of California, where the early work on this plasmid was conducted. Its colonies can be easily differentiated from colonies of other non-recombinant based colonies on the basis of their colour.
Structure of pUC19 Vector
pUC19 is similar to plasmid pBR322 in structure the only difference between the two is the single mutation and the lack of the rop gene which is responsible for its high copying nature . It has a double stranded circular DNA ,contains 2686 base pairs and has the weight of 1.75 x 10^6 Da.
pUC19 full form
UC= University of California
19= numerical designation
- It has been constructed using the ampicillin resistance gene and the pMB1 origin of replication from pBR322.
- The plasmids contains sites of various restriction enzymes. The more the number of restriction sites the more the production of desired fragments for cloning.
- pUC19 contains the lac operon of E.coli and the 5′-terminal part of the lacZ gene encoding for the N-terminal part of B-galactosidase.
- Some of the restriction sites on pUC19 vector are EcoR1, HindIII, BamH1 and many more.
Blue-White Screening Method for pUC19 Vector
Blue-White screening method is an efficient and rapid technique of identification of recombinant bacteria. The technique relies on the activity of B-galactosidase which occurs in E.coli, that cleaves lactose into glucose and galactose.
In this method a blue pigment is formed when B-galactosidase catalyzes hydrolysis of a synthetic substance known as X-gal. Due to X-gal hydrolysis it forms galactose and 5-bromo-4-chloro-3-hydroxy indole ,which undergoes dimerization and oxidation and forms a blue substance in the medium.
Therefore the cells which contain B-galactosidase activity forms blue colonies and the ones which does not form white colonies on the agar medium. The recombinant cells containing the newly inserted DNA lacks B-galactosidase and hence appear white.
Application of pUC19 Vector
The application of high copying vectors like pUC19 is increasing from time to time. The major role they play are in the field of molecular biology and genetic engineering due to their simplicity and cost-effectiveness. Some of their applications are as follows:
- Cloning vector are used in the transfer of foreign desireable DNA fragment into the host cell for our various purposes.
- They are used for the engineering of the E.coli bacteria for insulin production.
- Cloning Vector can also be used for studying the structure and function of various proteins.
- They can be used to identify mutation in regions of the DNA and also gene defects as well.
- PRINCIPLES OF GENE MANUPULATION AND GENOMICS BY PRIMROSE AND TWYMAN