SNP genotyping

Bulk samples of dried leaves or kernels from up to eight D_{step one} 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 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 .

Data of adult hereditary assortment

Hereditary diversity ranging from parental lines is reviewed with genome-wider SNP indicators profile equestriansingles from the dominant complement investigation, group investigation, and also by an effective pairwise genome inspect for polymorphism between your mothers of each and every inhabitants. To have facts, discover Most file 8.

Genetic map build

Hereditary maps had been constructed for every individual populace just like the explained earlier using CarthaGene called of individualized R programs. In the 1st action, statistically powerful scaffold charts were constructed with marker ranges regarding at the least ten cM. When you look at the a moment step, ework charts who has as numerous markers that one can, while maintaining a great LOD get >step 3.0 on the robustness off marker orders. In the end, the entire maps were acquired from the keeping more indicators using bin-mapping . CentiMorgan (cM) ranges was basically determined playing with Haldane's mapping setting . Private genetic charts and you can genotypic data employed for construction of one's maps (Even more file 4) have been transferred at MaizeGDB according to the endeavor phrase CORNFED .

Physical chart coordinates of SNPs

Chromosome and updates tasks regarding SNPs of one's MaizeSNP50 BeadChip offered by the manufacturer (Illumina ® , Hillcrest, Ca, USA), are based on this new B73 AGPv1 system with quite a few markers devoid of a beneficial chromosome and you can/otherwise standing suggestions. We ergo did an alternative mapping of your SNPs to the B73 AGPv2 construction playing with BWA . The new assignments were used for everyone analyses between the physical mapping pointers. Assignments can be found in A lot more document cuatro.

Offered an effective chromosome and also the relevant hereditary chart of an individual people, we computed the fresh marker positions towards B73 system. From the physical and you will genetic positions, i created an initial Marey chart with all the syntenic markers. So it Marey chart is actually smoothed using cubic spline interpolations , producing good 'bare' Marey map that was compelled to be monotonic. Upcoming places in which mapping advice is not having (for example, areas IBD about parents) was in fact masked, producing 'masked' Marey maps (Even more file nine). The fresh new detailed processes is actually said in A lot more file 8.