SNP genotyping

Bulk samples of dried leaves or kernels from up to eight D₁ plants derived from the same D₀, were used for DNA extraction using the cetyl trimethylammonium bromide (CTAB) procedure. DNA samples were adjusted to 50 to 70 ng/?l and 200 ng per sample were used for genotyping. DH line purity and integrity was first checked using a custom 96plex VeraCode assay (Illumina ® , San Diego, CA, USA) with genome-wide SNP markers to ensure that the lines carried only one of the parental alleles at each SNP, that they did not carry alleles of the inducer line and that they were derived https://datingranking.net/pl/amino-recenzja/ from true F₁ plants. For a subset of DH lines, 13 proprietary SNP markers assayed with the KASP™ technology (LGC Genomics, Berlin, Germany) were used for testing line purity and integrity. True DH lines were then used for genotyping with the Illumina ® MaizeSNP50 BeadChip on an Illumina ® iScan platform. Array hybridization and raw data processing were performed according to manufacturer’s instructions (Illumina ® ). Raw data were analyzed in Illumina ® ‘s Genome Studio software version v2011 (Illumina ® ) using an improved version of the public cluster file (MaizeSNP50_B.egt, ). SNP data were filtered based on the GTscore using a threshold of 0.7. Heterozygous SNPs were set to missing values (NA) and only markers with a minor allele frequency >0.1 per population were used for mapping. For each population, the allele of the central line was coded as the ‘A’ allele, and the allele of the founder line was coded as ‘B’ allele (Additional file 4). Raw genotyping data of parents and DH lines are available at NCBI Gene Expression Omnibus as dataset GSE50558 .

Investigation away from parental genetic assortment

Hereditary diversity anywhere between adult traces is examined that have genome-wide SNP indicators from the dominant complement analysis, party research, and also by a pairwise genome examine for polymorphism between the mothers of each people. To possess details, see Extra document 8.

Genetic chart framework

Hereditary maps was developed for every private population while the explained before playing with CarthaGene entitled of custom R texts. In the first action, statistically sturdy scaffold charts was designed with marker ranges of within minimum ten cM. Into the a moment step, ework maps that has had as many markers you could, while maintaining an effective LOD get >3.0 into the robustness out-of marker orders. In the long run, the whole maps was indeed gotten of the keeping a lot more indicators using bin-mapping . CentiMorgan (cM) ranges was calculated playing with Haldane’s mapping setting . Personal genetic maps and you may genotypic analysis employed for structure of one’s maps (A lot more file cuatro) was transferred at the MaizeGDB beneath the endeavor acronym CORNFED .

Physical map coordinates from SNPs

Chromosome and you may updates tasks off SNPs of your MaizeSNP50 BeadChip supplied by the product manufacturer (Illumina ® , San diego, California, USA), depend on the newest B73 AGPv1 assembly with several indicators lacking an excellent chromosome and/or updates recommendations. I therefore did a different mapping of your own SNPs on the B73 AGPv2 assembly using BWA . The fresh projects were used for everyone analyses between the bodily mapping suggestions. Projects can be found in More file 4.

Considering a great chromosome and relevant genetic map of men and women inhabitants, we calculated the latest marker positions into the B73 construction. Because of these bodily and you will hereditary positions, we built a first Marey chart that contains most of the syntenic markers. That it Marey chart is actually smoothed using cubic spline interpolations , generating an effective ‘bare’ Marey chart which had been compelled to be monotonic. After that regions in which mapping information are lacking (like, locations IBD throughout the parents) was basically masked, producing ‘masked’ Marey charts (More document 9). This new intricate techniques try said when you look at the More document 8.