Bacterial Artificial Chromosome

1. ‫م‬ْ‫ح‬َّ‫الر‬ ِ‫هللا‬ ِ‫م‬ ْ
‫س‬ِ‫ب‬
ِ‫ن‬
ِ‫م‬ْ‫ي‬ِ‫ح‬َّ‫الر‬

2. Presented by
Ali Zia Kamboh

3. Presented to
Ms Sana Hussain

4. Bacterial Artificial
Chromosome

5. Table of Contents
• Bacterial Artificial Chromosomes
• Cloning
• Components in Bacterial Artificial Chromosomes
• Mechanism
• Method
• Application

6. Cloning
• Bacterial Artificial Chromosomes(BAC) utilize a
cloning system derived from a plasmid found in
E.coli.
• Allows one to develop much larger pieces of DNA
compared to standard plasmids (10,000bp vs
350,000bp)
• Reduces numbers of clones need to sequence a
genome by 35x
• First used by Hiroaki Shizuya in 1992

7. Components in a bac
• RepE: for plasmid replication and
regulation of copy number
• Hind III and Bam HI: Sites of Cloning
• oriS: The origin of replication
• CmR: Chloramphenicol resistance gene,
selection tool
• ParA,ParB,ParC: the genes that regulate
the partitioning of plasmids to daughter
cells during division.

8. Mechanism
• Developed by taking parts of
of the F’ and turning it into a
vector.
• The origin of replication
allows the host cell to
recognize the new DNA
insert.
• Location of HindiII and
BAmHI within LacZ changes
X-gal metabolism when
target DNS is incorporated
• CmR induces antibiotic
resistance to target DNA
containing vectors

9. Vector and
donor DNA
both cut by
same
restriction
enzyme
Fragments
are mixed
and sticky
ends
hybridize via
H-bonds
DNA Ligase
seals gaps by
forming
phosphodiest
er linages

10. Method
• DNA Fragments of interest are
isolated and cleaved using
restriction enzymes
• The BAC is digested by restriction
enzymes around the cloning site
(HindlII and BamHI)
• Recombinant DNA is formed
(F’plasmid and target DNA) using
DNA ligase.
• New recombinant DNA is inserted
into compliant cells and plated

11. Method
• As bacterial cells grow and divide they also
amplify the BAC DNA which can be isolated
• CmR and LacZ distinguish between successful
transmission of target gene into bacterium.

12. Applications
• BACs allow cloning and maintenance of
large segments of DNA, making them
useful in whole genome mapping
• Independent Chromosome allows easy
isolation, and pure inserts.
• Allows genetic information to be stored
from organisms that aren’t easily grown in
cultures.
• Study of large pathogenic viruses

13. References
• W.C. Nierman, T.V. Feldblyum, in Encyclopedia of Genetics, 2001
• G.M. Weinstock, in Encyclopedia of Genetics, 2001
• “What Are BAC Libraries?” Facts, The Public Engagement Team at the Wellcome
Genome Campus, 25 Feb. 2015,.
• StudiousGuy. “Cloning Vectors: Types & Characteristics.” StudiousGuy, StudiousGuy,
19 Sept. 2018, studiousguy.com/cloning-vectors-types-characteristics/.
• Kevinshe. “THE BIG BAD BAC: BACTERIAL ARTIFICIAL CHROMOSOMES.” SCQ,
6 Sept. 2006, www.scq.ubc.ca/the-big-bad-bac-bacterial-artificial-chromosomes/.

14. Questions?

15. Thank You