Non‐Mendelian Single‐Nucleotide Polymorphism Inheritance and Atypical Meiotic Configurations are Prevalent in Hop

Abstract

Hop (Humulus lupulus L.) breeding programs seek to exploit genetic resources for bitter flavor, aroma, and disease resistance. However, these efforts have been thwarted by segregation distortion including female-biased sex ratios. To better understand the transmission genetics of hop, we genotyped 4512 worldwide accessions of hop, including cultivars, landraces, and over 100 wild accessions using a genotyping-by-sequencing (GBS) approach. From the resulting ~1.2 million single-nucleotide polymorphisms (SNPs), prequalified GBS markers were validated by inferences in population structures and phylogeny. Analysis of pseudo-testcross (Pt) mapping data from F 1 families revealed mixed patterns of Mendelian and non-Mendelian segregation. Three-dimensional (3D) cytogenetic analysis of late meiotic prophase nuclei from two wild and two cultivated hop revealed conspicuous and prevalent occurrences of multiple, atypical, nondisomic chromosome complexes including autosomes. We used genome-wide association studies (GWAS) and fixation index (F st ) analysis to demonstrate selection mapping of genetic loci for key traits including sex, bitter acids, and drought tolerance. Among the possible mechanisms underlying the observed segregation distortion from the genomic data analysis, the cytogenetic analysis points to meiotic chromosome behavior as one of the contributing factors. The findings shed light on long-standing questions on the unusual transmission genetics and phenotypic variation in hop, with major implications for breeding, cultivation, and the natural history of Humulus.

3D Molecular Cytology of Hop (Humulus lupulus) Meiotic Chromosomes Reveals Non-disomic Pairing and Segregation, Aneuploidy, and Genomic Structural Variation

“…Plants were collected and fixed according to previously described methods (Zhang et al, 2017 ), with exceptions on timing and paraformaldehyde concentration during the meiocyte buffer A (MBA) steps. Briefly, hop panicles were field collected and immediately fixed in Farmer's fluid (3:1 ethanol:acetic acid) overnight, replaced with Farmer's fluid for a second overnight period, and exchanged into 70% ethanol for storage at −20C.…”

“…Three-dimensional images were collected on a DeltaVision deconvolution microscope, using a 60X lens and 0.2 micron Z-step optical sections as described (Zhang et al, 2017 ) for DAPI imaging, but also including FITC, TRITC, or CY5 imaging for FISH probes that fluoresce green, red, or far-red, respectively. Distance measurements between the 5S rDNA FISH signals were obtained as point-to-point Euclidean distances using the Measure Distances program, manually selecting the brightest voxel centered in the X, Y, and Z dimensions for any given FISH signal.…”

“…The long history of cultivation includes colchicine-induced polyploidization (Roborgh, 1969 ) and the introgression of genetically distinct wild populations (Small, 1980 ; Reeves and Richards, 2011 ). Recent genetic, genomic, and quantitative trait analyses have demonstrated that the genome of hop is complex and structurally diverse (Zhang et al, 2017 ). Irregularities in hop transmission genetics are reflected in non-Mendelian segregation distortion and sex ratio bias (Seefelder et al, 2000 ; Jakse et al, 2008 , 2011 ; McAdam et al, 2013 , 2014 ).…”

“…Meiotic chromosome behavior is implicated as a cause for the unusual transmission genetics that are found in hop (Zhang et al, 2017 ). In sexually reproducing organisms, meiosis reduces the genomes from diploid to haploid (John, 1990 ) and homologous chromosomes from each parental genome undergo pairing, synapsis, recombination (John, 1990 ).…”

“…Among the hallmarks of meiosis that are cytologically evident are (1) the presence of a telomere bouquet in early prophase that guides homology search and subsequent synapsis, (2) the presence of bivalents at diakinesis during late prophase, (3) complete and equal separation of homologs at meiosis I, and (4) the production of 4 haploid nuclei with 1:1:1:1 segregation, as evidenced from tetrad analysis. For hop, non-disomic and heteromorphic sex chromosome figures have been noted and speculated to impact segregation patterns (Sinotô, 1929 ; Jacobsen, 1957 ; Neve, 1958 ; Haunold, 1991 ; Shephard et al, 2000 ; Zhang et al, 2017 ). However, neither the early prophase telomere bouquet nor the post-meiotic segregation has been characterized cytogenetically.…”