PRJNA181184
Sorghum bicolor Phenotype or Genotype
Digital Genotyping (DG) is a restriction enzyme targeted genome resequencing method for genetic analysis of sorghum and other grass species with large repeat-rich genomes. DG templates are generated using one of three methylation sensitive restriction enzymes that recognize a nested set of 4-, 6- or 8 bp GC-rich sequences, enabling varying depth of analysis and integration of results among assays. Use of methylation sensitive enzymes resulted in a 6-fold enrichment of unique versus repetitive DG sequences. FseI-depth analysis is particularly useful for DG marker-assisted breeding, diversity analysis, and genetic map construction. This enzyme recognizes the 8 bp sequence GGCCGGCC, present in or near many nuclear genes but absent in the high copy plastid genome. Variation in sequencing efficiency among DG markers was correlated with template GC-content and length. The expected DG allele sequence was obtained 97.3% of the time with a ratio of expected to alternative allele sequence acquisition of >20:1. A genetic map aligned to the sorghum genome sequence with an average resolution of 1.47 cM was constructed using 1772 DG markers from 137 recombinant inbred lines. The DG map enhanced the detection of QTL for variation in plant height and precisely aligned QTL such as Dw3 to underlying genes/alleles. Higher-resolution NgoMIV-based DG haplotypes were used to trace the origin of DNA on SBI-06, spanning Ma1 and Dw2 from progenitors to BTx623 and IS3620C. DG marker analysis identified the correct location of two miss-assembled regions and located seven super contigs in the sorghum reference genome sequence.