Bacteria as Hosts, Hosts Multiplication, Chimeric Vectors, Bacterial Strain, DNA Molecules, Gram Negative Bacteria, Pseudomonas, Rhizobium

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Chimeric DNA Molecular Probes and Gene Libraries Chimeric DNA Molecular Probes and Gene Libraries Introduction Restriction Enzymes for Cloning Technique of Restriction Mapping Restriction Clevage and Gel Electrophoresis Construction of a Restriction Map Use of Partial Digests end Labelling and Hybridization in Restriction Mapping Construction of Chimeric DNA Cloning of Plasmid Vectors Directional Cloning of DNA Fragments with Heterologus Protruding Termini Adding Poly dA at 3' Ends of DNA Clone Blunt End Ligation by T4 DNA Ligase Cloning in Phage λ and Cosmid Vectors Cloning in phage λ Vectors Cloning in Cosmid Vectors Hosts for Cloning Vectors Bacteria as Hosts Eukaryotes as Hosts Molecular Probes Preparation of Probes Genomic DNA Probes

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Bacteria as hosts

Bacteria are often used as hosts for multiplication of chimeric vectors or recombinant DNA molecules. Therefore, a bacterial strain should be available, which should be modified or selected for high transformation efficiency to be used as a host. Bacteria can be classified into. (i) gram negative bacteria e.g. Escherichia coli. Pseudomonas, Rhizobiumand (ii) gram positive bacteria e.g. Bacillus subtilis and Streptomycessp.

Ordinarily same plasmid can not maintain itself in stable form in both classes of bacteria and, therefore, plasmid v ctors for two classes usually differ. While gram negative bacteria can be used as hosts for plasmids, phages and' cosmid vectors, gram positive bacteria can be used as hosts for only plasmids. Chimeric vector is inserted into bacterial cells and transformed bacterial colonies are selected and used for multiplication in suspension cultures. The chimeric DNA is retrieved, whenever required.

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c DNA Probes Synthetic Oligonucleotides as Probes RNA Probes or Riboprobes Labelling of Probes Radiolablled Probes Non Radioactive Probes Biotin Labelled Probes Digoxigenin Labelled Probes Alternatives to Biotin and Digoxigenin Labelling Amplification of DNA Probe Signals Techniques Used in Molecular Probing Southern Nothern and Western Blotting Southern Blotting Northern Blotting Western Blotting Dot Blots and Slot Blots Applications of Molecular Probes Construction and Screening of Genomic and cDNA Libraries Genomic Library by Shotgun Experiment cDNA Library from mRNAs Colony or Plaque Hybridization for Screening of Libraries Chromosome Walking and Characterization of Chromosome Segments Chromosome Jumping or Hopping Libraries BAC Libraries and Assembly of BACs into Contigs

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