Pollen tetrad-based visual assay for meiotic recombination in Arabidopsis
- Kirk E. Francis,
- Sandy Y. Lam,
- Benjamin D. Harrison,
- Alexandra L. Bey,
- Luke E. Berchowitz, and
- Gregory P. Copenhaver*
- Department of Biology and Carolina Center for Genome Scientists, University of North Carolina, Chapel Hill, NC 27599
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Edited by Susan R. Wessler, University of Georgia, Athens, GA, and approved January 8, 2007 (received for review October 10, 2006)
Abstract
Recombination, in the form of cross-overs (COs) and gene conversion (GC), is a highly conserved feature of meiosis from fungi to mammals. Recombination helps ensure chromosome segregation and promotes allelic diversity. Lesions in the recombination machinery are often catastrophic for meiosis, resulting in sterility. We have developed a visual assay capable of detecting Cos and GCs and measuring CO interference in Arabidopsis thaliana. This flexible assay utilizes transgene constructs encoding pollen-expressed fluorescent proteins of three different colors in the qrt1 mutant background. By observing the segregation of the fluorescent alleles in 92,489 pollen tetrads, we demonstrate (i) a correlation between developmental position and CO frequency, (ii) a temperature dependence for CO frequency, (iii) the ability to detect meiotic GC events, and (iv) the ability to rapidly assess CO interference.
Footnotes
- *To whom correspondence should be addressed. E-mail: gcopenhaver{at}bio.unc.edu
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Author contributions: K.E.F., L.E.B., and G.P.C. designed research; K.E.F., S.Y.L., B.D.H., A.L.B., L.E.B., and G.P.C. performed research; K.E.F., L.E.B., and G.P.C. analyzed data; and G.P.C. wrote the paper.
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The authors declare no conflict of interest.
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This article is a PNAS direct submission.
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See Commentary on page 3673.
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This article contains supporting information online at www.pnas.org/cgi/content/full/0608936104/DC1.
- Abbreviations:
- CO,
- cross-over;
- GC,
- gene conversion;
- LMS-PCR,
- ligation-mediated suppression-PCR;
- FTL,
- fluorescent-tagged line;
- I1,
- interval 1;
- I3,
- interval 3;
- I5,
- interval 5;
- CFP,
- cyan fluorescent protein;
- ECFP,
- enhanced CFP;
- YFP,
- yellow fluorescent protein;
- EYFP,
- enhanced YFP;
- RF,
- recombination frequency;
- NPD,
- nonparental ditype;
- DCO,
- double cross-over.
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Freely available online through the PNAS open access option.
- © 2007 by The National Academy of Sciences of the USA
