Chromosome Jumping or Hopping Libraries, Specific Gene, Human Disorders, Huntington Disease, Duchenne Muscular Dystrophy

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Chromosome Jumping (or Hopping) Libraries

In most cases, we identify the genes by their protein products, which help in the isolation and cloning of a specific gene. However, for many human disorders, which are each controlled by a single gene (e.g. Huntington disease, cystic fibrosis, duchenne muscular dystrophy, etc.), the gene product is not known despite intensive investigations. Such genes are cloned by a process, in which the gene is identified primarily by its map position.

However, even if the distance of the gene from a molecular marker is as small as lcM, which is the limit of resolution, this will be equivalent to 100 to 5,000 kb. Therefore, if a segment carrying the gene is to be cloned using the linked molecular marker, such a segment carrying both the gene and the marker will be too large to be cloned.

In order to bring the molecular marker closer to the gene of interest, 'chromosome jumpping' approach has been utilized. The technique of 'chromosome jumping' is based on the following steps.

(i) depending upon the distance between the gene and the marker, decide about the distance of 'jumps' or 'hopsize' (e.g. 100 kb or 200 kb); (ii) genomic DNA molecules in the range of a particular size (say 80 kb-130 kb in case of 100 kb 'hopsize' or 160-240 kb for 'hopsize' of 200k'J) are selected through pulsed-field gel electrophoresis;

(iii) for circularization of DNA segments, ligation between two ends of each long linear DNA molecule was allowed using T4 ligase in the presence of supPF⁺;(iv) DNA circles obtained in step (iii) above are digested with EcoRI; v) the vector λCh3Δi1lac, an amber mutated phage vector (supF^-) is also cut with EcoRI and used for cloning small DNA fragments representing the junctions of the circularized genomic DNA molecules and carrying supP;

(vi) the cloned DNA fragments obtained in step (v) above represent the jumping library, which can be plated on a bacterial host and screened through the technique of plaque hybridization described earlier.

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Steps involved in chromosome jumping

Steps Involved in Chromosome Jumping

Steps Involved in chromosome Jumping

Steps Involved in Chromosome Jumping

The above technique of chromosome jumping will help narrowing the gap between the gene and available molecular markers. After several cycles of chromosome jumping followed by cloning the regions that are closer to the gene, it will be possible to approach very close to the desired gene and clone it.

It is thus abvious that using the technique of 'chromosome jumping' it will be possible to isolate, clone and characterize genes, whose gene products are unknown. The gene for cystic fibrosis was isolated using this technique. It could also be shown that 'cystic fibrosis' disease is caused by a single base substitution in this gene.

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