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Standard XII
Biology
Ligase
Name the five...
Question
Name the five key tools for accomplishing the tasks of recombinant DNA technology. Also mention the functions of each tool.
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Solution
Introduction :
The five key tools involved in recombinant DNA technology are as follows:
1.Enzymes involved in DNA manipulation
2.Cloning vectors
3.Gel electrophoresis
4.PCR
5.Host organism
Enzymes involved in DNA manipulation:
Enzymes involved in DNA manipulation are as follows:
1.Restriction enzymes
These enzymes serve as molecular scissors to cut DNA into defined fragments.
These enzymes recognize special palindromic nucleotide sequences of 4-8 nucleotides in length in DNA.
Restriction endomucelases (a type of restriction enzyme) are used to isolate gene of interest and to create sticky ends in plasmid vectors.
Example - Eco R I
2.Polymerase enzyme
These enzymes cause extension of DNA strands in the 5’ to 3’ direction by using template DNA.
E.g., Taq polymerase in PCR
3.Ligase enzyme
The DNA ligase joins two DNA strands by forming a phosphodiester bond between them.
It is known as molecular glue.
It is used to join genes of interest with the cloning vector.
E.g., T4 DNA ligase is used in rDNA technology.
Cloning vector :
Cloning vehicles that carry the gene of interest into the recipient cell are called cloning vectors.
Different types of cloning vectors are listed below:
Plasmids and viruses are commonly used as vectors.
Plasmids are extra chromosomal circular DNA present in prokaryotic organisms such as bacteria, that are capable of replicating themselves.
The most commonly used vectors are plasmids such as pBR322 and pUC18.
Ti plasmid obtained from Agrobacterium is the commonly used vector for plants.
Disarmed retroviral vectors are commonly used for animals.
Gel electrophoresis:
Gel electrophoresis is a technique used to separate DNA fragments based on their sizes under the influence of an electric field.
PCR:
Polymerase Chain Reaction is used to amplify the gene of interest.
Using PCR even small samples of DNA can be amplified and used in rDNA technology.
It involves three steps - denaturation, annealing and extension.
Host organism :
The organism in which the gene of interest (with or without vector) is inserted and multiplied is known as the host organism.
Escherichia coli is commonly used for multiplication of various genes.
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Ligase
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