Question

compare and contrast 8 different types of cloning vectors.

1. prokaryote bacterial plasmid

2. Yeast Eukaryotic plasmid

3. Bacteriophage Lambda

4. Cosmos

5. Bacteriophage P1

6 PACs

7 BACs

8 YACs

Answer 1

Cosmids

Analysis of eukaryotic genes and the genome organisation of eukaryotes requires vectors with a larger capacity of cloned DNA than plasmids or $\lambda$ lambda.

Cosmids use the $\lambda$ packaging system to package large DNA fragments bounded by $\lambda$ cos site, which circularize and replicate as plasmids after infection of E.coli cells. Some cosmid vectors have two cos sites, and are cleaved to produce two cos ends, which are ligated to the ends of target fragments and packaged into lambda particles. Cosmids have a capacity for cloned of 30-45 kb.

YAC Vectors: Yeast artificial chromosomes can be constructed by ligating the components required for replication and segregation of natural yeast chromosomes to very large fragments of target DNA, which may be more than 1 Mb in length. YAC vectors contain two telomeric sequences (TEL), one centromere (CEN), one autonomously replicating sequence (ARS) and genes which can act as selectable markers in yeast.

BAC Vectors: Bacterial artificial chromosomes are based on the F factor of E. coli and can be used to clone up to 350 kb of genomic DNA in a conveniently handled E.coli host. They are a more stable and easier to use alternative to YACs.

In case of organisms with small genome sizes, such as E.coli, a genomic library could be constructed in a plasmid vector as only 5000 clones (of average insert size 5 kb) would give a greater than 99% chance of cloning the entire genome (4.6 x 10⁶ bp). Most libraries from organisms with larger genomes are constructed using phage $\lambda$ , cosmid, bacterial artificial chromosome (BAC) or yeast artificial chromosome (YAC) vectors. Thse accept inserts of approximately 23, 45, 350 and 1000 kb respectively, and thus fewer recombinants are needed for complete genome coverage than if plasmids were used.