Predicting the Size of the Progeny Mapping Population Required to Positionally Clone a Gene

Abstract

A key frustration during positional gene cloning (map-based cloning) is that the size of the progeny mapping population is difficult to predict, because the meiotic recombination frequency varies along chromosomes. We describe a detailed methodology to improve this prediction using rice (Oryza sativa L.) as a model system. We derived and/or validated, then fine-tuned, equations that estimate the mapping population size by comparing these theoretical estimates to 41 successful positional cloning attempts. We then used each validated equation to test whether neighborhood meiotic recombination frequencies extracted from a reference RFLP map can help researchers predict the mapping population size. We developed a meiotic recombination frequency map (MRFM) for $1400 marker intervals in rice and anchored each published allele onto an interval on this map. We show that neighborhood recombination frequencies (R-map, .280-kb segments) extracted from the MRFM, in conjunction with the validated formulas, better predicted the mapping population size than the genome-wide average recombination frequency (R-avg), with improved results whether the recombination frequency was calculated as genes/cM or kb/cM. Our results offer a detailed road map for better predicting mapping population size in diverse eukaryotes, but useful predictions will require robust recombination frequency maps based on sampling more progeny.

High Resolution Genetic and Physical Mapping of a Major Powdery Mildew Resistance Locus in Barley

“…One gene belongs to the RLK and the rest to the NBS-LLR gene family making each of them a potential candidate gene for the MlLa-H locus. The observed physical to genetic distance ratio at the MlLa-H locus (∼1.16 Mb/cM) and physical distance of flanking markers would require screening of an additional 6,000 meioses (based on the formula of Dinka et al, 2007) to provide a chance of observing any additional recombination events between the cluster of the remaining co-segregating markers and the resistance gene to provide genetic evidence for rejecting several of the found candidate genes. The possibility of resolving the correct candidate gene by genetics and recombination, however, remains a theoretical option, since the candidate genes were observed in a susceptible genotype.…”

Genome-wide Association Studies in Maize: Praise and Stargaze

“…Only a few QTLs are generally detected via linkage mapping in each experiment. Further fine mapping of QTL to a more narrowly precise genetic position and cloning of the underlying gene, as large secondary populations are generally required to achieve sufficient map resolution (Dinka et al, 2007), are particularly resource-and time-consuming processes. The large and complex maize genome, more than 85% of which consists of repetitive sequences (Schnable et al, 2009) further slows progress in QTL fine mapping and cloning.…”

Development of a next-generation NIL library in Arabidopsis thaliana for dissecting complex traits

“…This was accomplished using a modest population size of BC1S3 plants (n = 286) and the interval could be narrowed down further through genotyping at a higher number of loci and increasing the population size [56]. Regardless, in a six-week period we have identified a population encompassing recombinants in the 4.8 Mb region identified as causal during the QTL validation experiment, translating to a 3-fold change in total time versus a breeding scheme utilizing inbred NILs.…”