Vector and it's properties , types

Vector is the DNA molecules that acts as a carrier for recombinant technology...

1. Vectors Introduction Properties of good vector Types of vector Plasmid  vector Cosmids Phagemids Phage vector or Bacteriophage vector Shuttle vector

2. Vector It is a  biological tool  in rDNA technology. It is used for the delivery of desired foreign  DNA  into the host cell. Vectors are the DNA molecules, which can carry a foreign DNA fragment to be cloned. They are selfreplicating in an appropriate host cell. The most important vectors are plasmids, bacteriophages, cosmids and phasmids .

3. C apable of replicating inside the host. C ompatible restriction site for insertion of DNA molecule (insert). Capable of autonomous replication inside the host ( ori site). Smaller in size with low molecular weight and able to incorporate larger insert size. Have a selectable marker for screening of recombinant organism . Characteristics Of An Ideal Vectors

4. Classification of vectors Based on the nature and sources, the vectors are categorized into following classes: 1 . Plasmids 2 . Bacteriophages 3 . Cosmids 4 . Phagemids 5 . Shuttle vectors

5. Plasmid Plasmids are extrachromosomal , double stranded , circular, self-replicating DNA molecules. almost all the bacteria have plasmids containing a low copy number (1-4 per cell) or a high copy number (10-100 per cell). The size of plasmids varies from 1 to 500 kb. Usually plasmids contribute to about 0.5 to 5.0% of the t otal DNA of bacteria. Examples of plasmid vectors are: pBR322 pUC 18/19

6. Plasmids have the following advantages as cloning vehicle 1.It can be readily isolated from the cells. 2.It possesses a single restriction site for one or more restriction enzymes. 3. Insertion of foreign DNA does not alter the replication properties. 4.It can be reintroduced into cell. 5.Selective marker is present. 6. Transformants can be selected easily by using selective medium. 7 . Multiple copy numbers are present in a cell.

7. pBR322 vector :- It is one of the first vectors to be developed by   Boliver  and  Rodriguez  in 1977.

8. The name pBR322 denotes the following: P- plasmid B- Boliver R- Rodriguez 322- Differentiate it from the other plasmid produced in the same laboratory E.g. –  pBR325 ,  pBR327 , etc. It is 4363 base pair long. It carries two sets of antibiotic resistance gene: Ampicilli Tetracycline It contains only single or unique recognition site for 12 different restriction enzymes: Pst I Sac I Pvu I

9. pUC 18/19 It is derivative of pBR322. Much smaller (~ 2.7kb). Ampicillin resistance gene ( bla ). ColE1  origin of replication. It is 2,686bp long. The name pUC18/19 derived from: P- Plasmid. UC- university of California.

10. BACTERIOPHAGES Bacteriophages or simply phages are the viruses that replicate within the bacteria. ln case of certain phages, their DNA gets incorporated into the bacterial chromosome and remains there permanently . Phage vectors can accept short fragments of foreign DNA into their genomes. The advantage with phages is that they can take up Iarger DNA segments than plasmids. Hence phage vectors are preferred for working with genomes of human cells.

11. On the basis of structure bacteria phases are of two types: Head and Tail Phages- E.g.:  lambda phage. Filamentous phage- E.g.:  M13 phage. Lambda phage vectors Its genome size is about 48,502 bp. It contains origin of replication, genes for head and tail protein and enzymes for  DNA replication It has more than one recognition sequence for almost all the restriction enzymes. It should be larger than 38 kb and smaller than 52 kb to packaged into phage particles.

12. M13 phage vectors M13 vectors are used to obtain single- stranded copies of cloned DNA. It is 6407 nucleotides long. It is circular and 6.4kb in size . It is used to produce several copies of M13 mp series of vectors. Example-  M13mp8, M13mp9  etc.

13. Cosmids vector It is a type of  hybrid plasmid . It contains  lambda phage cos sequence . Cosmids = cos sites + plasmid. Genomic size of cosmids is about 30 to 52 kb. If they have suitable origin of replication than they can replicate as Plasmid within the host cells, E.g.-  SV40 Ori , ColE1 ori . It also contains selectable marker such as Ampicillin resistance gene. Collins  and  Hohn  in 1978 was first to described cosmids .

14. Phagemid Vector A plasmid vector which contains an origin of replication from a phage, in addition to that of the plasmid, is termed as Phagemids . pBlue script SK (+/-)  is a type of phagemid vector of 2,958 bp derived from the pUC19. It consists of the following: ColE1 ori Phage f1 (M13) origin of replication. A small portion of lacZ gene. MCS within lacZ Gene from Lac promoter. Phage T7 and T3 promoter sequence. Ampicillin gene for Ampicillin resistance.

15. Shuttle vector Shuttle vectors are created to replicates in cell of different type of species. They contain two origin of replication, in which one is particular for each host species, also those genes required for their replication and not provided by the host cell. This type of vectors are developed by recombinant techniques.