What is a lambda DNA?
Lambda DNA, a linear, double-stranded phage DNA containing 12 bp single-stranded complementary 5′-ends, is derived from an Escherichia coli bacteriophage (Bacteriophage lambda cI857 Sam7).
Why is lambda DNA used in restriction digestion?
Since the lambda genome is approximately 48,000 base pairs, each restriction enzyme will cut the DNA several times and generate restriction fragments of different sizes. In this activity, three separate samples of lambda DNA will be cut using three different restriction enzymes, and one sample will remain undigested.
What is DNA restriction analysis?
The DNA restriction analysis experiment demonstrates that DNA can be precisely manipulated and that it behaves as predicted by the Watson-Crick structure. Students use restriction enzymes, the scissors of molecular biologists, to cut DNA from the bacteriophage lambda.
What are lambda vectors?
Bacteriophage lambda has been in use as a cloning vector for over 25 years, and has been used extensively as an expression vector. The efficiency of packaging and infection, and the simplicity of plaque screening are advantages of lambda as a cloning vector.
Who discovered lambda DNA?
Lambda was originally discovered in 1951 by Esther Lederberg (1951) at the University of Wisconsin (Madison), when she serendipitously found it was released from the laboratory Escherichia coli strain K-12 after ultraviolet irradiation.
What is the size of the lambda genome?
48,490 bp
Bacteriophage lambda has a 48,490 bp genome that contains two 5′ overhangs to circularize the phage genome once it has entered a bacterial cytoplasm.
What is restriction analysis used for?
Restriction analysis is to identify restriction mapping sites in DNA sequences using appropriate enzyme sets and enzyme filtering criteria as per specific experimental requirements.
Why is a lambda phage used in cloning?
Bacteriophages such as lambda phages can also be used as effective vectors for transferring recombinant DNA molecules into cells for cloning. A lambda phage is a bacteriophage that infects E. Coli cells. This phage, like any virus, readily undergoes two types of life cycles – the lytic cycle and the lysogenic cycle.
What are lambda insertion vectors?
Figure 01: Lambda Phage Insertion vector consists of a single recognition site. The primary function of this vector is to develop cDNA libraries derived from eukaryotic mRNA sequences. Moreover, it can only accommodate lengths of foreign DNA ranging in between 05-11 kb.
What is the function of lambda restriction enzyme?
This enzyme is encoded by Lambda itself and acts like a restriction enzyme during the replication of the phage DNA. It is an endonuclease specific for the cos-site in multimeric phage DNA. The ends of the resulting monomeric DNA (called cos ends) are similar to the sticky (or cohesive) ends produced by common restriction enzymes.
Why are some lambda DNA fragments circular?
When the digest is not heated before the gel run (lane 1), a number of b and c fragments have joined to form band a, which is in size the sum of b and c. It is very likely that a number of the Lambda DNA molecules was circular (because of the cos-site formed) already before the digestion.
What is the function of the enzyme terminase in lambda DNA?
note: The 12 nucleotide 5’ overhangs at the cos-ends of the linear Lambda DNA are the result of a cut by the enzyme terminase. This enzyme is encoded by Lambda itself and acts like a restriction enzyme during the replication of the phage DNA. It is an endonuclease specific for the cos-site in multimeric phage DNA.
Why does lambda DNA digest show Extra bands after electrophoresis?
After electrophoresis, the gel band pattern of a Lambda DNA digest may show an extra band, due to the joining (and forming the cos-site) of the two end fragments. If this happens (it will always, to some extent..), the bands containing the separate end fragments will be present in an amount lower than calculated..