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

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.

journal_name

Nucleic Acids Res

journal_title

Nucleic acids research

authors

Anand R,Villasante A,Tyler-Smith C

doi

10.1093/nar/17.9.3425

subject

Has Abstract

pub_date

1989-05-11 00:00:00

pages

3425-33

issue

9

eissn

0305-1048

issn

1362-4962

journal_volume

17

pub_type

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