Integrated cytogenetic map of mitotic metaphase chromosome 9 of maize: resolution, sensitivity, and banding paint development

Abstract

To study the correlation of the sequence positions on the physical DNA finger print contig (FPC) map and cytogenetic maps of pachytene and somatic maize chromosomes, sequences located along the chromosome 9 FPC map approximately every 10 Mb were selected to place on maize chromosomes using fluorescent in situ hybridization (FISH). The probes were produced as pooled polymerase chain reaction products based on sequences of genetic markers or repeat-free portions of mapped bacterial artificial chromosome (BAC) clones. Fifteen probes were visualized on chromosome 9. The cytological positions of most sequences correspond on the pachytene, somatic, and FPC maps except some probes at the pericentromeric regions. Because of unequal condensation of mitotic metaphase chromosomes, being lower at pericentromeric regions and higher in the arms, probe positions are displaced to the distal ends of both arms. The axial resolution of FISH on somatic chromosome 9 varied from 3.3 to 8.2 Mb, which is 12-30 times lower than on pachytene chromosomes. The probe collection can be used as chromosomal landmarks or as a "banding paint" for the physical mapping of sequences including transgenes and BAC clones and for studying chromosomal rearrangements.

Karyotype and Identification of All Homoeologous Chromosomes of AllopolyploidBrassica napusand Its Diploid Progenitors

“…These results suggested that an ancient chromosome rearrangement occurred between the two duplicated regions of C4B in either the A or C genomes after the diversification from a common ancestor. The future of plant molecular cytogenetics is promising in terms of both methods and applications (Kato et al 2005;Walling et al 2005;Lamb and Birchler 2006;Danilova and Birchler 2008;Guerra 2008;Pires and Hertweck 2008;Lysak et al 2010;Xiong et al 2010). The robust novel chromosomal painting technique developed here will assist the understanding of chromosome pairing, homoeologous recombination, and genome evolution in the genus Brassica and will facilitate new applied breeding technologies in this genus that rely upon identification of individual chromosomes.…”

Single-copy gene fluorescence in situ hybridization and genome analysis: Acc-2 loci mark evolutionary chromosomal rearrangements in wheat

“…The FISH resolution on condensed mitotic chromosomes is not very high. The axial resolution of FISH on maize somatic chromosome is 3-8 Mb, but it is possible to distinguish separate FISH signals that are about 200 kb apart (Danilova and Birchler 2008). Double Acc-2 FISH signals were observed on chromosomes 3A, 3D, 3B, and 4A of bread wheat; 4A t and 3G of T. timopheevii; and 3D of Ae.…”

“…For genomes enriched with repeats, a repeat-free probe for a particular genic region either can be produced from cDNA or developed from genomic DNA by sequence analysis and PCR amplification of the repeat-free region, and used in direct FISH as was shown in maize (Wang et al 2006;Lamb et al 2007;Danilova and Birchler 2008) and barley (Ma et al 2010). In direct FISH, fluorochromes are incorporated directly into DNA probes and the procedure does not need detection step.…”

Improvements in cytological preparations for fluorescent in situ hybridization in Passiflora

“…However, they require long incubation periods, up to 2 h or more with some species (De Melo and Guerra, 2003;Marcon et al, 2005). Aiming to achieve a faster or more efficient results, other enzymes have been used, such as macerozyme (Fukui and Nakayama, 1996), pectolyase and cellulase Onozuka RS (Danilova and Birchler, 2008;Yuan and Tomita, 2009), Ultrazym (Almeida and Carvalho, 2004), and Pectinex (Viccini et al, 2006;Guimarães et al, 2009), among others.…”