Human and other mammalian genomes are very large, therefore, vectors are needed to clone large DNA fragments. Many vectors have been prepared that aid in the characterisation and expression of large genes or gene complexes. These vectors exhibit varied cloning capacity (table). Table Maximum DNA Insert Possible with Different Cloning Vectors Vectors Hosts Insert Size
Vectors for Cloning Larger DNA Fragments Read More »
Cloning vector is a DNA molecule in which when a foreign DNA is integrated achicves the capability to replicate it within itself to produce many clones of recombinant DNA. Characteristics of a Cloning Vector Its size should be small. It should bear the capability to self-replicate within the host cell. It should carry a restriction
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The target DNA or the DNA insert isolated and enzymatically cleaved by the selective restriction endonuclease, are ligated (joined) through the action of ligase enzyme to the vector DNA. As a result, an rDNA molecule known as cloning-vector-insert DNA construct is produced. Direct Ligation of Cohesive Termini (DNA Ligase) The target gene can be inserted
Insertion of Target DNA Into a Vector Read More »
In recombinant DNA (rDNA/rec DNA) technology, DNA molecules from two different sources are joined and inserted into a host organism to get products useful for human use (figure). A clone refers to a group of cells or organisms obtained from one progenitor, thus indicating that the members of a clone are genetically same. This is
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The process of developing proteins having desired functions by manipulating their stability and specificity is termed protein engineering. This can be done by taking two main approaches, rational design and irrational design (or directed evolution). In the former case, knowledge of the protein structure and function is considered and a rational gene mutation is planned.
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Genetic engineering involves deliberate DNA manipulation in organisms to alter their genes. Although the organisms whose genes are being altered may not be microbes, but the substances and techniques involved are obtained from microbes and adapted for use in more complex organisms. Historical Background The term genetic engineering initially was used for various techniques used
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The industrial applications or therapeutic uses of enzymes/proteins can be appropriately brought into use by increasing their half-lives or thermostability. Proteins with enhanced stability can be obtained by the following methods: Addition of disulfide bonds, Changing asparagine to other amino acids, Reducing the free sulfhydryl groups, Single amino acid changes, and Improving kinetic properties of
Increasing the Stability and Biological Activity of Proteins Read More »
Baeyer’s strain theory successfully explained the relative stability of cyclic compounds having 6-membered rings (or more appropriately having 5-membered rings), but it failed to explain the stability of higher compounds. Since the valency deviation (d) of seven, eight. etc., membered rings is significantly high, these compounds should also be unstable; but the stability of these
Sachse Mohr’s Theory (Theory of Strainless Rings) Read More »
A carboxylic acid with a long, unbranched, saturated or unsaturated aliphatic tail (or chain) is called a fatty acid. Natural fatty acids found abundantly carry carbon atoms in even number because they are biosynthesised using acetyl-CoA (a co-enzyme having a two carbon atom group). Hydrolysis of the ester linkages in fat or biological oil (both
Classification of Fatty Acids Read More »
If electron-withdrawing groups are present on the benzene ring of benzoic acid, the benzoate ion stabilises and acidity thus increases. This is attributed to inductive and resonance effects. In case of p-nitrobenzoic acid and its m-isomer, based on the inductive effect alone the m-isomer would be assumed to be stronger than the p-isomer but the reverse is also true.
Effect of Substituents on Acidity Read More »