Question

On which plasmid are Bacterial Artificial Chromosome (BAC) vectors based? Enlist the important sequences present in these vectors.

Hint:

The low copy number ($1-2$ copies per cell) of BAC vectors is their key advantage: it prevents toxic overproduction of cloned gene products and reduces recombination-mediated deletions of large DNA inserts.

Answer 1

Step 1: Concept
Bacterial Artificial Chromosomes (BACs) are high-capacity cloning vectors designed to clone very large segments of DNA (up to 300 kb) in E. coli cells.

Step 2: Plasmid Base of BAC Vectors
BAC vectors are based on the naturally occurring F-plasmid (Fertility plasmid) of E. coli. The F-plasmid replication is strictly controlled, maintaining a low copy number of only $1-2$ copies per cell, which ensures stable cloning of large genomic inserts without rearrangement.

Step 3: Important Sequences Present in BAC Vectors
BAC vectors contain several essential sequence motifs:
1. $oriS$ and $repE$: Direct the unidirectional replication of the BAC plasmid.
2. $parA$, $parB$, and $parC$: Active partitioning genes that ensure the stable segregation of plasmids into daughter cells during cell division.
3. Selectable Marker: Usually a chloramphenicol resistance gene ($cm^R$) to select for transformants.
4. Cloning Site: A multiple cloning site (MCS) containing unique restriction sites, often embedded within a reporter gene like $lacZ$ for blue-white screening.
5. Cos and LoxP sites: Optional sites added for selective cleavage or recombination.
Final Answer: BAC vectors are based on the F-plasmid of E. coli. Their essential sequences include replication control genes ($oriS$, $repE$), partitioning genes ($parA$, $parB$, $parC$), a chloramphenicol resistance marker ($cm^R$), and a multiple cloning site (MCS).