The maternal Chrysanthemum horaimontanum (2n=18) and the paternal Tanacetum vulgare (2n=18) were artificially crossed and produced hybrid strains which were raised by embryo rescued and axenic cultured in vitro and later acclimatized in vivo. The hybrid plants had the chromosome number of 2n=18, 19 and 27. The technique of genomic in situ hybridization (GISH) made it possible to distinguish between the two parental genomes in the chromosome complement of the F₁ hybrid plants. Some of the hybrid plants showed genome rearrangement as a translocation in certain somatic chromosomes of C. horaimontanum and T. vulgare. Parental chromosomes that carried the 45S rDNA could be marked by simultaneous bicolor hybridization using the digoxigenin labelled total genomic DNA of T. vulgare probe and the biotin labeled pTa71 probe. A signal of 45S rDNA was hybridized in the nine chromosomes of C. horaimontanum, while three signals were hybridized in the nine chromosomes of T. vulgare in the chromosome complement of their F₁ hybrid.

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The maternal Chrysanthemum horaimontanum (2n=18) and the paternal Nipponanthemum nipponicum (2n=18) were artificially crossed and produced F₁ hybrid plants. GISH was applied on meiotic chromosomes in pollen mother cells (PMCs) of the F₁ hybrid to study chromosome organization and segregation of the two parents. Uni- Bi- and multivalents were observed during diakinasis showed only single terminal chiasmata. Multi-valents were observed during meiosis-I and showed nonhomologous pairing occurred between homoeologous chromosomes among the same haploid chromosome set and between chromosomes of the two parents. The most common meiotic abnormalities were those related to the irregular segregation of univalent chromosomes, characterized by laggards, fragments and multiple bridges in anaphase-telophase I and telophase II. In general, these chromosomes gave rise to micronuclei during telophase that remained until the second meiosis. GISH displayed that all chromosomal bridges were related to N. nipponicum genome. Tetrads showed different sizes of nuclei and elimination of one of the parental genome from others. One of the very interesting phenomenon of parental genome separation and spatial organization that was detected for the first time in the pollen grains of the intergeneric F₁ hybrid between C. horaimontanum and that of N. nipponicum the two genomes were differentially stained and fluoresced as distinct domains of two different colors following GISH.

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Molecular phylogeny of Ochrosia sensu lato (Ochrosia sensu stricto and Neisosperma) was investigated by maximum parsimony analysis of sequence data from the internal transcribed spacer (ITS) of nuclear ribosomal DNA. Twenty-one species including two varieties (25 accessions), were defined as the ingroup. Three outgroup species were selected from tribe Vinceae (Kopsia arborea, K. flavida, and Rauvolfia serpentina). The ITS sequence data showed that Ochrosia s.l. is a highly defined monophyletic group (supported with 100% bootstrap value). As shown in the strict consensus tree, all Neisosperma species were placed at the basal part of the tree with N. glomerata and N. nakaiana as the basalmost taxa. All species belong to Ochrosia s.str. were grouped into one clade and nested within Neisosperma resulting in monophyletic Ochrosia s.str. and paraphyletic Neisosperma. This suggested that Neisosperma should be included into Ochrosia, and the two groups are treated as a single genus. The analysis also showed that subdivision of Ochrosia s.l. into sections based on fruit morphology (fibrous vs. massive and thick endocarp) is not supported.

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Molecular phylogeny of Ochrosia sensu lato (Ochrosia sensu stricto and Neisosperma) was investigated by maximum parsimony analysis of sequence data from chloroplast DNA (rps16 intron) and rps16 intron + Internal Transcribed Spacer (ITS). Nineteen species including two varieties (23 accessions) were defined as the ingroup, comprising ten species of Ochrosia s.str. and nine species of Neisosperma. For analysis using rps16 intron region, Rauvolfia sumatrana, R. verticillata and R. serpentina, Vinca major, and V. minor of the tribe Vinceae were used as the outgroup. The aims of this study are to evaluate the usefulness of rps16 intron to infer phylogenetic relationships within Ochrosia sensu lato and to confirm conclusions from previous analysis. In this study, rps16 region was proved to be sufficiently informative to infer phylogenetic relationships at infrageneric level. The strict consensus of most parsimonious trees produced from rps16 sequences data showed that Ochrosia species formed a monophyletic clade highly supported with 97% bootstrap value, whilst Neisosperma was paraphyletic. The Ochrosia clade was found to be nested within Neisosperma, with N.acuminata as a sister taxon to it. All Ochrosia and Neisosperma species were grouped into one clade. However, this clade was not well supported (only has 61% bootstrap value). Consequently, although the results does not contradict recognizing only one genus (Ochrosia sensu lato), but it is not convincing enough. The strict consensus tree produced from sequence data analysis using rps16 intron + ITS showed a more robust monophyletic clade of Ochrosia sensu lato, supported with 100% bootstrap value. Neisosperma - again - was found to be paraphyletic, with monophyletic clade of Ochrosia (strongly supported with 93% bootstrap value) nested within, suggesting the inclusion of Neisosperma into Ochrosia and recognition of only one genus. These findings supported our previous result, using ITS region.

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A phylogenetic analysis of Ochrosia sensu lato (Ochrosia sensu stricto and Neisosperma) was performed based on 14 macromorphological characters scored from 19 species including two varieties (23 accessions), and three outgroup from genus Rauvolfia, i.e. R. sumatrana, R. verticillata and R. serpentina. For the same species, this data was analyzed together with molecular dataset from our previous studies. The result of total datasets analysis (of ITS, rps16 intron and macromorphological characters) confirmed the monophyly of Ochrosia sensu lato. Ochrosia sensu stricto appeared to be monophyletic. The clade nested within paraphyletic Neisosperma and together they form a strongly supported monophyletic clade of Ochrosia sensu lato. Most of the morphological characters used in this study were found to be homoplasious. The existence of fibrous-fruited group and cavity-fruited group was not supported, thus subdivision of Ochrosia into sections (or subgenera) based on this character as proposed by several authors cannot be applied.

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The internal transcribed spacer (ITS) region of 18S-26S nuclear ribosomal DNA (nrDNA) in seven species and one variety of Pinguicula (Lentibulariaceae) in Cuba has been sequenced. Although the eight taxa have been taxonomically divided into two subgenera such as Isoloba and Pinguicula with the other species grown in the other regions, and further they are divided into three sections such as Agnata, Discoradix and Homophyllum. The taxa of Pinguicula in Cuba formed a single clade, and it was not consistent with the current taxonomy based on morphology. Six taxa of Pinguicula in Cuba studied have been classified in section Agnata together with Mexican Pinguicula agnata and P. ibarrae, however, they were not ITS-phylogenetically, fully or partially supported by the data. Although the Cuban taxa of Pinguicula were quite different in morphology and life form from each other, they were ITS-phylogenetically close relatives to each other. The species of Pinguicula in Cuba were isolated phytogeographically and ITS-phylogenetically from those in the Southeastern United States and Mexico.

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