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Double-strand breaks on artificial chromosomes in yeast
L O Ross1, D Zenvirth, A R Jardim
1Department of Molecular Microbiology, Tufts University, Boston, MA 02111, USA.
Chromosoma
|September 1, 2000
Summary
Yeast artificial chromosomes show low meiotic crossing over. Adding yeast DNA sequences restored recombination, demonstrating that double-strand break sites on the insert dictate recombination activity.
Area of Science:
- Genetics
- Molecular Biology
- Yeast Biology
Background:
- Yeast artificial chromosomes (YACs) made of bacteriophage DNA have limited meiotic crossing over compared to natural yeast chromosomes.
- Natural yeast chromosome recombination is initiated by DNA double-strand breaks (DSBs).
Purpose of the Study:
- To investigate the relationship between DSB sites and recombination in YACs.
- To understand how specific DNA sequences influence YAC recombination properties.
Main Methods:
- Physical analysis of YACs containing a 12.5 kb insert from yeast chromosome VIII.
- Mapping of DSB sites on artificial and natural yeast chromosomes.
- Deletion analysis of specific DSB sites within the YAC insert.
Main Results:
- DSBs were infrequent on YACs, except within the 12.5 kb chromosome VIII insert.
- DSB positions on the YAC insert precisely correlated with known DSB sites on chromosome VIII.
- Deleting two specific DSB sites from the insert reduced DSBs and crossing over.
Conclusions:
- The recombination behavior of YACs is strongly influenced by the presence and characteristics of inserted yeast DNA sequences.
- DSB sites within inserted DNA segments are critical determinants of meiotic recombination frequency and localization on artificial chromosomes.