Probing the W chromosome of the codling moth, Cydia pomonella, with sequences from microdissected sex chromatin

Abstract

The W chromosome of the codling moth, Cydia pomonella, like that of most Lepidoptera species, is heterochromatic and forms a female-specific sex chromatin body in somatic cells. We collected chromatin samples by laser microdissection from euchromatin and W-chromatin bodies. DNA from the samples was amplified by degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR) and used to prepare painting probes and start an analysis of the W-chromosome sequence composition. With fluorescence in situ hybridization (FISH), the euchromatin probe labelled all chromosomes, whereas the W-chromatin DNA proved to be a highly specific W-chromosome painting probe. For sequence analysis, DOP-PCR-generated DNA fragments were cloned, sequenced, and tested by Southern hybridization. We recovered single-copy and low-copy W-specific sequences, a sequence that was located only in the W and the Z chromosome, multi-copy sequences that were enriched in the W chromosome but occurred also elsewhere, and ubiquitous multi-copy sequences. Three of the multi-copy sequences were recognized as derived from hitherto unknown retrotransposons. The results show that our approach is feasible and that the W-chromosome composition of C. pomonella is not principally different from that of Bombyx mori or from that of Y chromosomes of several species with an XY sex-determining mechanism. The W chromosome has attracted repetitive sequences during evolution but also contains unique sequences.

Holocentric chromosomes: convergent evolution, meiotic adaptations, and genomic analysis

“…1c). Chromosomes can be broken by X-ray irradiation (Hughes-Schrader and Schrader 1961), or more precisely by laser dissection (Fuková et al 2007). Chromosomal fragments must persist after breakage, and micronuclei resulting from a failure to segregate chromosome fragments should not be observed.…”

Sex chromosome composition revealed in Characidium fishes (Characiformes: Crenuchidae) by molecular cytogenetic methods

“…gomesi did not reveal any similarity among themselves or with any sequences deposited in GenBank to date (Table 3). This kind of dead-end result has been reported before when studying sequences present in the sex chromosomes of Leporinus elongatus (Nakayama et al 1994;Parisi-Maltempi et al 2007), Parodon hilarii Reinhardt, 1866 (Vicente et al 2003), Cydia pomonella L., 1758 (Fuková et al 2007) and Oryzias hubbsi Roberts, 1998 (Takehana et al 2012), for example. Of the seven distinct fragments analysed, only CgW9 evidenced similarity (∼73%) to a DNA transposon (Helitron-2 Dr) isolated from the genome of Danio rerio.…”

2D and 3D Chromosome Painting in Malaria Mosquitoes

“…Previously, the technique required the use of specialized microdissection needles that had to be manually controlled by an experienced user 28 . The advancement of laser capture microdissection (LCM) has resulted in a simplified tool better suited for isolating single cells 29,30 or individual chromosomes [31][32][33] with a lessened risk of contamination. This approach allows the user to study the genetic heterogeneities and chromosomal abnormalities that occur in single cells, instead of a consensus landscape that results from pooling multiple cells together [34][35][36] .…”