cDNA Library from mRNAs, Complementry DNA, Synthesiizing Proteins, leaves in Plants, Ovaries, Reticulocytes Mammals

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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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cDNA library from mRNAs

Complementary DNA (cDNA) libraries can also be prepared by isolating mRNAs from tissues which are actively sythesizing proteins like roots and leaves in plants ovaries or reticulocytes in mammals. etc. The mRNA is used for copying it into cDNA through the use of reverse transcriptase.

A cDNA molecule can be made double stranded and cloned and discussed later in Chapter 8. However cDNA clones will differ from genomic clones in lacking the introns present in split genes, but may have the advantage of being capable to be expressed in bacteria which do not have the machinery to process HnRNA (obtained from split genes) into mRNA.

Synthesis of cDNA from mRNA, using reverse transcripts enzyme

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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