Toward a Molecular Cytogenetic Map for Cultivated Sunflower (Helianthus annuus L.) by Landed BAC/BIBAC Clones

Abstract

Conventional karyotypes and various genetic linkage maps have been established in sunflower (Helianthus annuus L., 2n = 34). However, the relationship between linkage groups and individual chromosomes of sunflower remains unknown and has considerable relevance for the sunflower research community. Recently, a set of linkage group-specific bacterial /binary bacterial artificial chromosome (BAC/BIBAC) clones was identified from two complementary BAC and BIBAC libraries constructed for cultivated sunflower cv. HA89. In the present study, we used these linkage group-specific clones (∼100 kb in size) as probes to in situ hybridize to HA89 mitotic chromosomes at metaphase using the BAC- fluorescence in situ hybridization (FISH) technique. Because a characteristic of the sunflower genome is the abundance of repetitive DNA sequences, a high ratio of blocking DNA to probe DNA was applied to hybridization reactions to minimize the background noise. As a result, all sunflower chromosomes were anchored by one or two BAC/BIBAC clones with specific FISH signals. FISH analysis based on tandem repetitive sequences, such as rRNA genes, has been previously reported; however, the BAC-FISH technique developed here using restriction fragment length polymorphism (RFLP)−derived BAC/BIBAC clones as probes to apply genome-wide analysis is new for sunflower. As chromosome-specific cytogenetic markers, the selected BAC/BIBAC clones that encompass the 17 linkage groups provide a valuable tool for identifying sunflower cytogenetic stocks (such as trisomics) and tracking alien chromosomes in interspecific crosses. This work also demonstrates the potential of using a large-insert DNA library for the development of molecular cytogenetic resources.

Fluorescence chromosome banding and FISH mapping in perennial ryegrass, Lolium perenne L.

“…This left the chromosomes well-spread and completely clean on the glass slide without any cytoplasmic debris. An additional advantage of the well-spread and debris-free chromosome preparation has been the enhanced sensitivity of FISH signals [30, 31]. Here, we have successfully mapped a small target sequence, LpGI , onto a specific chromosome band of L. perenne using both genomic DNA and cDNA probes.…”

Sunflower Hybrid Breeding: From Markers to Genomic Selection

“…Using the newly developed 25 K SNP array in sunflower Livaja et al (2016) were able to construct a linkage map based on 6,355 SNP markers for the RIL population NDBLOSsel × CM625. The connection between genetic linkage maps and the sunflower karyotype was finally made by developing a molecular cytogenetic map for H. annuus ( Feng et al, 2013 ). BAC and BIBAC clones with known genetic locations were used in fluorescence in situ hybridization (FISH) experiments to address the individual chromosomes.…”

Integration of Lupinus angustifolius L. (narrow-leafed lupin) genome maps and comparative mapping within legumes

“…Genetically anchored BAC clones were used both to identify individual chromosomes in metaphase spreads and to establish a complete FISH-based karyotyping cocktail that permitted simultaneous identification of all 20 soybean chromosome pairs (Findley et al 2010 ). Genome maps that associate all the chromosomes to corresponding linkage groups have been constructed for a number of plant species (Feng et al 2013 ) including Medicago truncatula (Schnabel et al 2003 ), Lotus japonicus (Ohmido et al 2010 ), and P. vulgaris (Fonseca et al 2010 ).…”