Lambda-based vectors

Lambda-Based Vectors and Main Approaches to Constrnct... 

There are many different modifications to construct libraries however, the three ways to construct genomic libraries are the most commonly used and will be exemplified below using genomic lambda-based vectors (Figure 1.2-3, [12-14]). 

Christensen, A. 2001. Bacteriophage lambda-based expression vectors. Molecular Biotechnology 17, 219-224. 

In this section, common vectors used for everyday cloning and subcloning like plasmid and M13 vectors will not be discussed. Only vectors suitable for cloning relatively large DNA inserts will be mentioned with the exception of lambda phage-based vectors specially designed to clone cDNA. 

Libraries constructed in lambda-based, BAG, and PAG vectors are widely used now in different laboratories, and almost nobody is currently working with libraries constructed in YAG vectors. 

Armed with this new tool, Schena et al. (1996) created a microarray of 1,046 human cDNAs of unknown sequence. They were derived from human peripheral blood lymphocyfes fransformed wifh Epsfein-Barr virus. Suitably sized inserts [>600 base pairs (bp)] were cloned into a lambda vector, subsequently infected into an Escherichia coli strain, and finally amplified by polymerase chain reaction (PCR) using 5 -amino-modified primers. The resulting 5 -amino-modified cDNA amplicons were then arrayed onto sily-lated microscope slides. Next, the expression levels in human Jurkat cells undergoing heat shock or phorbol ester induction were examined. 

Researchers have expanded upon the natural diversity generated by the immune system. In 1989 a novel vector system based on the bacteriophage lambda was used to express a synthetic combinatorial library of Fab fragments of the mouse antibody repertoire in Escherichia coli [22]. This system allowed rapid and easy identification of monoclonal Fab fragments that bind a given antigen, in a form suitable for genetic manipulation. For example. 

Supplementary features of some binary vectors include overdrive and T-DNA transfer stimulator sequences (to improve T-strand synthesis), and a cos region from lambda bacteriophage to allow maintenance of cosmid-based plant genomic libraries in AgrobacTeriutn (51 and refs contained therein). 

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