Chromosome Number and Karyotype Variation in Codiaeum variegatum Cultivars

Codiaeum variegatum (L.) Blume is one of the most popular ornamental foliage plants. It encompasses more than 300 recognized cultivars valued by their wide range of leaf shapes and vivid foliage colors. Thus far, only limited information is available regarding the genetic basis of their leaf morphological variation. This study investigated the chromosome numbers and karyotypes of seven phenotypically diverse cultivars. Root-tip cells were fixed, mounted, and observed under light microscopy. Results showed that chromosome numbers in the mitotic metaphase of the seven cultivars were high and variable and ranged from 2n = 66, 70, 72, 76, 80, 82, 84, to 2n = 96, indicating that the cultivars are polyploid and some could be aneuploid. Genetic mosaics occurred in one of the seven cultivars. Additionally, each cultivar had its own karyotype. There were no relationships between chromosome numbers or karyotypes and leaf morphology. Results from this study suggest that the morphological diversity among cultivars of this species could be in part attributed to high variation in chromosome numbers and karyotypes. Codiaeum variegatum (L.) Blume, commonly known as crotons, are small evergreen trees and shrubs that are popular as ornamental plants because of their attractive foliage. Currently, more than 300 croton cultivars are reported in the world (Brown, 1995), and they are produced as either landscape plants or containerized foliage plants (Chen et al., 2005). The foliage varies widely in leaf size, shape, color, and variegation pattern among the cultivars. Leaf sizes range from small (6 cm long · 1 cm wide) to large (35 cm long · 13 cm wide), and margins can be entire or trifurcate. Leaf shapes differ from simple ovate to linear to spiral; some are slightly or deeply cut, connected along the blade only by the midrib. Because they are prized by the leaf morphology, The Croton Society (2009) categorized croton cultivars into nine types based on their leaf shapes: 1) broad leaf; 2) oak leaf; 3) semioak-leaf; 4) spiral leaf; 5) narrow leaf; 6) very narrow leaf; 7) small leaf; 8) interrupted leaf; and 9) recurved leaf. Native to the Mollucan Islands of Indonesia (Brown, 1995), crotons were first introduced to England in 1804, and hybrids were developed in Belgium and France during the 1800s. The hybrids were introduced into the United States in 1871 and subsequently were further hybridized in south Florida during the 1920s and 1930s, which resulted in the release of more than 70 hybrid cultivars, commonly known as Florida hybrids (Brown, 1995). Crotons are monoecious with poor seed set. Seed propagation often results in plants with different phenotypes (Sharma and Bal, 1958). Thus, once a hybrid cultivar is selected, it is propagated through stem cuttings (Chen and Stamps, 2006). Although crotons have a long history of introduction and hybridization, there is limited information regarding the genetic basis of their morphological variation. Sharma and Bal (1958) examined chromosome numbers in root-tip cells of 24 croton varieties collected from India and found that all the varieties studied were polyploid. Subsequent research by Pancho and Hilario (1963) from the Philippines and Chennaveeraiah and Wagley (1985) from India also suggested that the croton cultivars they studied were polyploid. Recently, Ogunwenmo et al. (2007) studied root-tip cells of six crotons cultivated in Nigeria and reported that four of the six cultivars had 2n = 60, but one had 2n = 24 and the other 2n = 30. Florida has a rich history in croton cultivar development and also leads the nation in croton production (Brown, 1995). However, there is no information regarding the cytology of crotons cultivated in Florida. The objectives of this study were to examine the chromosome number and karyotype of selected crotons cultivated in Florida and to determine if any relationships exit between chromosome numbers or karyotype and leaf morphology. Materials and Methods Plant materials. Seven croton cultivars (Curly Boy, Gold Dust, Ice Tone, Mother and Daughter, Ralph Davis Hybrid, Yellow Petra, and Picasso’s Paint Bush; Fig. 1) were gathered from germplasm maintained at the University of Florida’s Mid-Florida Research and Education Center and from ornamental foliage plant nurseries in central and southern Florida. The seven cultivars represent five leaf types: spiral (‘Curly Boy’), small (‘Gold Dust’ and ‘Ice Tone’), interrupted (‘Mother and Daughter), broad (‘Ralph Davis Hybrid’ and ‘Yellow Petra’), and very narrow leaf (‘Picasso’s Paint Bush’) and are also among the most popular ones cultivated in Florida. Stem cuttings were rooted in 20-cmdiameter plastic pots using a sphagnum peatbased medium (Canadian peat: vermiculite: perlite at a 3:1:1 ratio based on volume), five pots per cultivar. After rooting, plants were grown in a shaded greenhouse under daily maximum photosynthetically photon flux (PPF) density of 350 mmol m s at a temperature range of 20 to 32 C. All plants were fertilized with a top-dress application of 8 g Multicote (15N–7P2O5–15K2O) controlledrelease fertilizer with an 8-month longevity (Haifa Chemicals Ltd., Haifa Bay, Israel) and hand-watered two to three times a week as needed. Six months after growing in the shaded greenhouse, the cultivars were used for the following cytological studies. Chromosome number determination. Ten root tips, two per pot, were collected from each cultivar in the morning and pretreated as described by Dyer (1979) with modifications. Briefly, root tips were treated with 2.0 mM 8hydroxyquinoline solution for 3 h at room temperature. The root tips were then fixed in a 1:3 acetic acid:absolute ethanol solution overnight at 4 C and preserved in a 70% ethanol solution at –20 C. Fixed root tips were softened by hydrolyzation in 1 M HCl solution at 60 C for 5 min, rinsed briefly in distilled water, and stained with 2% acetoorcein for 15 min. The root tips were mounted on a glass slide with a fresh drop of the stain and covered with a coverslip. The glass slide was warmed by alcohol flame for a few seconds. The coverslip was tapped gently with fine forceps and squashed using the thumb. Chromosome numbers were determined by counting at least 10 complete well-spread mitotic metaphase cells, one cell per root tip, using a Nikon Optiphot Pol microscope (Nikon Nippon Kogaku K.K., Tokyo, Japan). Images were taken by a Canon S3 IS digital camera (Cannon U.S.A., Inc., Lake Success, NY). Karyotype determination. Karyotypes were analyzed in reference to the method of Sharma and Bal (1958). At least four complete well-spread mitotic metaphase cells of each cultivar were photographed and the lengths of chromosome arms were measured. Chromosomes were classified into two types: A) chromosomes without secondary constrictions; and B) chromosomes with one to three secondary constrictions. According to the position of the primary constriction (centromere), Received for publication 19 Jan. 2010. Accepted for publication 22 Feb. 2010. We thank Green Terra, Inc., Coral Springs, FL, and Hermann Engelmann Greenhouses, Inc., Apopka, FL, for providing plant materials used in this study and Russell D. Caldwell for assistance in plant collection and for critical review of the manuscript. Current address: School of Ecological Technology and Engineering, Shanghai Institute of Technology, Shanghai, China. To whom reprint requests should be addressed; e-mail [email protected]. 538 HORTSCIENCE VOL. 45(4) APRIL 2010 type A chromosomes were divided into two subtypes: A1) metacentric to submetacentric chromosome (Stebbins, 1971) with centromere located at the middle or submiddle of the chromosome; and A2) acrocentric chromosome with centromere close to the terminal area. Type B chromosomes were divided into four subtypes based on the secondary constriction positions: B1) metacentric to submetacentric chromosome with a secondary constriction on the longer arm that divided it into equal parts; B2) metacentric to submetacentric chromosome with a secondary constriction on the longer arm that divided it into unequal parts; B3) acrocentric chromosome with a secondary constriction close to the end of the longer arm that divided it into unequal parts; and B4) chromosome with more than one secondary constriction. The final karyotype of each cultivar and/or the different karyotypes within a cultivar were the result of separate examinations of four well-spread mitotic metaphase cells.