Molecular cytogenetic study on the scleractinian coral Micromussaamakusensis (Veron, 1990) (Hexacorallia, Anthozoa, Cnidaria): isolation of five fluorescence in situ hybridization markers.

Scleractinian (stony) corals are foundational to reef ecosystems, yet their taxonomy remains unresolved due to morphological plasticity and limited cytogenetic data. This study presents the first molecular cytogenetic characterization of the scleractinian coral Micromussaamakusensis (Veron, 1990), employing fluorescence in situ hybridization (FISH) to isolate and map five DNA markers. Using the conventional Giemsa staining technique, M.amakusensis was found to have a diploid karyotype of 2n = 28, with a prominent homogeneously staining region (HSR) on the long arm of chromosome 12. Subsequently, five FISH markers designated as MA-H3 for histone H3, MA-5S for 5S rRNA, MA-18/28S for 18S-28S rRNA, MA-13C for centromeric region, and MA-TEL for telomeric region were cloned, sequenced, and mapped using FISH. FISH analysis revealed that the MA-H3 localized to the centromeric region of chromosome 1, MA-5S to the telomeric region of chromosome 4, MA-18/28S to the terminal region of chromosome 12 (coinciding with the HSR), MA-13C to the centromere of chromosome 13, and MA-TEL to multiple telomeric regions across several chromosomes. Sequence analysis confirmed marker identities and revealed conserved and novel repetitive elements. Furthermore, Genomic DNA hybridization (GDH) of whole-sperm DNA revealed signals collected at several telomeric regions, suggesting the presence of repetitive sequences. These cytogenetic markers enable the identification of at least 3 out of 14 chromosome pairs, allow for more precise karyotyping, and highlight chromosomal features that may help resolve coral classification and improve understanding of genome evolution. This research demonstrates the utility of molecular cytogenetics in stony coral systematics and provides new FISH markers for future comparative genomic studies.