Isolation of Genes,Ribosomal RNA,Phenotypic Genes,Regulatory Function,Promoter Gene,Xenopus,Centrifugation,Multiple Copies

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Isolation, Sequencing and Synthesis of Genes Isolation, Sequencing and Synthesis of Genes - Introduction Isolation of Genes Isolation of Genes Coding for Specific Proteins Isolation of Genes with Known or Unknown Products having Tissue Specific Expression Isolation of Genes with Known or Unknown Products Using DNA or RNA Probes Use of Heterologus Probes Use of Synthetic Probes Isolation of Genes Coding Use of Transposable Elements Transposon Tagging T-DNA-Insertion Mutagenesis for Isolation of Plant Genes Promoter Enhancer and Gene Trap for Isolation of Genes Differential Screeing and Differential Display Technique for Isolation of Genes Subtractive Hybridizatioon for Gene Isolation Map Based Cloning for Gene Isolation Isolation of Novel Genes by Asis Data at JNU New Delhi

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Your Ad Here	Isolation of Genes During the last three decades (1970-2000), significant progress has been made in the techniques for isolation of a variety of genes, including those for (i) ribosomal RNA; (ii) specific protein products; (iii) phenotypic traits with unknown product and those for (iv) regulatory function e.g. promoter genes, etc. Different techniques have been used for the isolation of these different types of genes and will be briefly described in this section.
Early attempts for isolation of ribosomal RNA genes in Xenopus As we know, ribosomes consist of ribosomal RNA (rRNA)( and proteins. Ribosomal RNA (rRNA) can be of four sizes namely, 28S, 16S, 5.8S and 5S (S strands for Svedberg units, a measure of speed with which a molecule sediments on centrifugation). This ribosomal RNA makes 80 per cent of cellular RNA and is synthesized from ribosomal genes, which could be isolated. Isolation of ribosomal genes was considered convenient due to their following three characteristic features:
(i) availability of homogeneous rRNA; (ii) difference between ribosomal RNA genes and other genes, due to relatively high G+C content in rRNA (rRNA has 45-60% G+C content in rRNA (rRNA has only 40% G+C); (iii) ribosomal genes are present in multiple copies, their number within a cell sometimes approaching several thousands In view of the above, ribosomal RNA genes were isolated for the first time (in 1965) in an amphibian named Xenopus, by Hugh Wallace and Max. L. Birnstiel, then working at the Univeristy of Edinburgh (U.K). Following steps were involved in this isolation: (i) rRNA was isolated from ribosomes of Xenopus and made radioactively labelled, due to its replication in a medium containing tritiated uridine; (ii) ribosomal DNA was isolated by density gradient centrifugation followed by its denaturation (G+C content of rDNA differs form that of bulk DNA and helps in it separation by centrifugation)	Your Ad Here
(iii) single stranded DNA was fixed on a filter paper carrying single stranded DNA; (v) DNA-RNA hybridization was allowed to take place; (vi) excess labelled RNA was washed; (vii) radioactivity was measured and duplex hybrids isolated, which on denaturation, gave single stranded DNA, which could be made double stranded Isolation of ribosomal DNA allowed the first characterization of ribosomal genes, which was followed by the isolation of 5S genes by Donald Brown. For a detailed account of isolation of ribosomal genes in Xenopus as originally done, readers are referred to an article Isolation of Genes in Scientific American (August. 1973)

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Sequencing of a Gene or a DNA Fragment Maxam and Gilbert's Chemical Degradation Method Sanger's Dideoxynucleotide Synthetic Method Automatic DNA Sequencers Slab Gel Versus Capillary Sequencers Slab Gel Sequencing Systems Capillary Gel Sequencing Systems Direct DNA Sequencing Using PCR Ligation Mediated PCR LMPCR DNA Sequencing Through Transcription DNA Sequencing by Hybridization Using Microarrays on DNA Chips DNA Sequencing by DE-MALDI-TOF Mass Spectrometry Synthesis of Genes Synthesis of Gene for Yeast Alanyl tRNA Synthesis of Oligonucleotides Synthesis of Three Duplex Fragments Synthesis of Gene from Three Duplex Fragments Synthesis of Gene for a True Precursor tRNA Total Synthesis of a Human Leukocyte Interferon Gene Gene Synthesis Machines Protection Chemistry and Amidites Permanent Protecting Groups Temporary Protecting Groups Synthesis of Gene Using PCR Synthesis of Genes From mRNA

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