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Home >>Molecular Probes and Gene Libraries >> Chromosome Walking and Characteristics of Chromosome Segments
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Chromosome Walking and Characteristics of Chromosome Segments - When a probe is used for the identification of a gene sequence in a genomic library, the probe may hybridize with a number of clones, each carrying a part of a large gene fragmented during preparation of genome library.

If we obtain partial digests (by digesting the DNA only partially) from the genome, different genomes (from large number of cells) may give fragments which have overlapping sequences, because sites cleaved in different genomes of the same organism, will differ being random.

Since none of these fragments may have entire sequence represented in the probe, overlapping sequences may be used to construct the original genomic sequence. Identification of fragments with overlapping sequences may be a key to the reconstruction or characterization of large chromosome regions. This is achieved by the technique popularly described as chromosome walking.

The technique of chromosome walking involves the following steps:
(i) from the genomic library select a clone of interest (identified by a probe) and subclone a small fragment from one end of the done (there is a technique available to subclone a fragment form the end);

(ii) the subcloned fragment of the selected clone may be hybridized with other clones in the library and a second clone hybridizing with the subclone of the first clone is identified due to presence of overlapping region;

(iii) the end of the second clone is then subcloned and used for hybridization with other clones to identify a third clone having overlapping region with the subcloned end of the second clone;

(iv) the third clone identified as above is also subcloned and hybridized with clones in the same manner and the procedure may be continued;

(v) restriction map of each selected clone may be prepared and compared to know the region of overlapping, So that identification of few overlapping restriction sites will amount to walking along the chromosome, or along a long chromosome segment.

Regions of chromosomes approaching 1000kb have been mapped following the above technique. Restriction maps of entire chromosomes can be prepared in this manner following the technique of chromosome walking.
 

 
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