Paper on Helianthus annuus (2n=34) BAC in situ. The BACs have a high content of retrotransposons, but Feng et al. from North Dakota use blocking DNA to prevent distributed hybridization signal, and identify BAC/BIBAC clones hybridizing specifically to each of the 17 chromosomes.
Feng, J., Liu, Z., Cai, X., & Jan, C.-C. (2013). Toward a molecular cytogenetic map for cultivated sunflower (helianthus annuus l.) by landed BAC/BIBAC clones. G3: Genes – Genomes – Genetics, 3 (1), 31-40.
Toward a Molecular Cytogenetic Map for Cultivated Sunflower (Helianthus annuus L.) by Landed BAC/BIBAC Clones
Jiuhuan Feng, Zhao Liu, Xiwen Cai and Chao-Chien Jan
USDA-Agricultural Research Service, Northern Crop Science Laboratory, Fargo, North Dakota
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.
International link to paper: http://www.g3journal.org/content/3/1/31.full.pdf+html
Local link to paper: HelianthusBACinsitu