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Chapter 31 : An Ordered Collection of Bacillus subtilis DNA Segments in Yeast Artificial Chromosomes

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An Ordered Collection of Bacillus subtilis DNA Segments in Yeast Artificial Chromosomes, Page 1 of 2

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Abstract:

A collection of chromosomal DNA fragments inserted in yeast artificial chromosomes (YACs) is represented in this chapter. YACs were obtained by random cloning of DNA, partially digested with ERI, in the YAC4 vector. To order the YACs, we first used 66 previously cloned genes as hybridization probes. Next, the ends of the inserts carrying these genes were isolated. The ends were used as probes to identify clones with overlapping inserts. Iteration of the procedure resulted in the ordered collection. The identity of several segments was ascertained by transformation of mutants. Despite the relatively high redundancy of the random collection, four gaps remain in the ordered collection. Maximum gap size was estimated by hybridizing DNA cleaved with several restriction enzymes with the ends of segments bordering the gaps. The ordered collection should be useful for genetic studies in .

Citation: Serror P, Azevedo V, Ehrlich S. 1993. An Ordered Collection of Bacillus subtilis DNA Segments in Yeast Artificial Chromosomes, p 473-474. In Sonenshein A, Hoch J, Losick R (ed), and Other Gram-Positive Bacteria. ASM Press, Washington, DC. doi: 10.1128/9781555818388.ch31

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Chromosomal DNA
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Pulsed-Field Gel Electrophoresis
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Figure 1

The ordered YAC clones containing DNA. The segments carried in individual YAC clones are represented by horizontal lines. They are placed according to their relationships with the neighboring clones and the genes they contain, but the extent of overlap between the segments has not been determined. YAC accession numbers are also mentioned. genes which hybridized with the corresponding YACs are indicated. Genetic position (in degrees) is given for and loci. Stars indicate markers which were identified in YAC clones by transformation of mutants. Filled and open circles represent left and right ends of inserts, respectively, which were (i) successfully isolated and (ii) shown to hybridize with the overlapping insert (except for inserts bordering the gaps). We failed to isolate one end of the insert carried on YAC 12-5 (carrying the gene), possibly because of their toxicity for and aberrant amplification in vitro. One end of the insert from YAC 15-37 (close to ), signalled by a vertical line, hybridized with the insert carried by YAC 15-634 (close to ), which shows that it is composed of nonadjacent DNA segments. However, the neighboring clone, YAC 15-10, is likely to contain the segment adjacent to The positions of four gaps present in the collection are indicated by vertical arrows and letters G1 to G4. The numbers in parentheses refer to gap size.

Citation: Serror P, Azevedo V, Ehrlich S. 1993. An Ordered Collection of Bacillus subtilis DNA Segments in Yeast Artificial Chromosomes, p 473-474. In Sonenshein A, Hoch J, Losick R (ed), and Other Gram-Positive Bacteria. ASM Press, Washington, DC. doi: 10.1128/9781555818388.ch31
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References

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