Construction of yeast artificial chromosome libraries with large inserts using fractionation by pulsed-field gel electrophoresis - PubMed (original) (raw)

Construction of yeast artificial chromosome libraries with large inserts using fractionation by pulsed-field gel electrophoresis

R Anand et al. Nucleic Acids Res. 1989.

Free PMC article

Abstract

A method for constructing yeast artificial chromosome (YAC) libraries with large insert sizes is reported. High molecular weight human DNA was partially digested with EcoRI and cloned in the vector pYAC4. When unfractionated DNA was used, the mean YAC size was 120kb. Fractionation by pulsed-field gel electrophoresis using a 'waltzer' apparatus to remove small DNA fragments increased the mean YAC size to congruent to 220kb or congruent to 370kb depending on the fractionation conditions. Ligated DNA prepared by this method was stable at 4 degrees C and routinely yielded transformation efficiencies of greater than 700 colonies/micrograms. It should be possible to extend the method to produce even larger inserts and to use high molecular weight DNA from any source.

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References

    1. Cell. 1984 May;37(1):67-75 - PubMed
    1. Science. 1987 May 15;236(4803):806-12 - PubMed
    1. Cell. 1987 Jul 31;50(3):509-17 - PubMed
    1. Nucleic Acids Res. 1987 Aug 11;15(15):5925-43 - PubMed
    1. Nature. 1988 Sep 8;335(6186):184-6 - PubMed

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