prev  page PLNT3140 Introductory Cytogenetics
Lecture 17, part 2 of 4
next page

III.GENOMIC LIBRARIES

A library is a random set of clones, in which genomic sequences are represented (in the ideal) by a Poisson distribution within the library. In other words, because cloning is a largely random process, some sequences will be cloned several times, and others with be cloned very rarely. Consequently, it is necessary to use the Clark & Carbon formula to determine how many clones are necessary to be sure, within a certain probability, of encountering each sequence at least once. Some sequences which are hard to clone may be statistically underrepresented in the library.

A. DETERMINING THE NUMBER OF CLONES NEEDED


 As illustrated at right, if you have only a few clones, they are likely to be from different parts of the genome. As you keep drawing clones from the library, more and more sites in the genome are represented.  Because clones are chosen at random, some parts of the genome will be overrepresented, while for other parts of the genome, no clones will have been chosen. Finally, if you choose a large enough number of clones,  you can be sure that every part of every chromosome is represented in at least one of the clones.  A genomic library is a population of clones, each containing a unique fragment of genomic DNA, which together, represent the entire genome.

The following equation [Clark & Carbon (1976) Cell 9:91] allows us to calculate the number of genomic clones necessary to construct a genomic library:
 

where

B.Cloning in BACs

She K (2003) So you want to work with giants: the BAC vector. BioTech Journal 1:69-74.
First - Why not clone in YACs?


BAC vectors


The term "BAC" stands for Bacterial Artificial Chromosome, but it important to remember that these are prokaryotic artificial chromosomes, that is they are designed to replicate in bacteria, not in eukaryotic cells. While BACs are actually derived from the E. coli F' plasmid, BACs are distinct from ordinary plasmids by having a number of features to optimize the ability to work with large inserts.

Working with large inserts
Unless otherwise cited or referenced, all content on this page is licensed under the Creative Commons License Attribution Share-Alike 2.5 Canada
prev  page PLNT3140 Introductory Cytogenetics
Lecture 17, part 2 of 4
next page