University of Nebraska - Lincoln DigitalCommons@University of Nebraska - Lincoln Agronomy & Horticulture -- Faculty Publications Agronomy and Horticulture Department 1984 Identification of Chromosomes with Secondary Constrictions in Melilotus Species S. E. Schlarbaum Kansas State University L. B. Johnson United States Department of Agriculture Herman J. Gorz United States Department of Agriculture Francis A. Haskins University of Nebraska-Lincoln, [email protected] Follow this and additional works at: https://digitalcommons.unl.edu/agronomyfacpub Part of the Plant Sciences Commons Schlarbaum, S. E.; Johnson, L. B.; Gorz, Herman J.; and Haskins, Francis A., "Identification of Chromosomes with Secondary Constrictions in Melilotus Species" (1984). Agronomy & Horticulture -- Faculty Publications. 307. https://digitalcommons.unl.edu/agronomyfacpub/307 This Article is brought to you for free and open access by the Agronomy and Horticulture Department at DigitalCommons@University of Nebraska - Lincoln. It has been accepted for inclusion in Agronomy & Horticulture -- Faculty Publications by an authorized administrator of DigitalCommons@University of Nebraska - Lincoln. The Journal of Heredity 75:23-26. 1984. Identification of chromosomes with secondary constrictions in Me/ilotuB species ABSTRACT: Secondary constrictions were determined in one chromosome pair in the com­ plements of M. intesta, M. macrocarpa, M. italica (subgenus Micromeli/otus), and M. alba and M. officinalis (subgenus Meli/otus). The karyotypes of M. intesta and M. macrocarpa were found to be similar. Chromosomes of M. italica were larger than the chromosomes of the other four Melilotus species. The chromosome size in M. italica suggests the pres­ ence of large chromosomes in an ancestral or pro-Melilotus prototype. The chromosomes with satellites of M. intesta and M. macrocarpa appear to differ in morphology from the satellite chromosomes of M. alba and M. officinalis by a paracentric inversion. The mor­ phology of the satellite chromosomes in M. italica is thought to rep resent a more primitive type than in the other Meli/otus species studied . S. E. Schla r baum T HE SW EETCLOVER GEN US, Melilotu s and M . italica. Addit ionally, a comparison Ada ns., contains forage and nitrogen fixing was made between the satellite chromosomes LB. Johnson species of agricultural importance. Taxo­ of these species and the satellite chromosomes of M . alba and M . officinalis for evidence of H.J.Gorz nomically, Melilot us is divided into two subgenera 10, (E ll )Melilotus Schulz and Mi­ kar yotypic evolution. F.A. Haskin s cromelilotus Schulz. Isely' considers the subgenus Melilotus to consist of nine biennial Materials and Methods species, including the commercially important M . alba Desr. and M. officinalis (L.) Larn., Seed of each species was obtained from the while the subgenus Mi crom elilotus contains collection of Melilotus germplasm that is kept II annual species, fewof which have economic in cold storage at the University of Nebraska. value as crops!". Other taxonomists, most M . infest a (Bdn. 61-98) and M . macrocarpa notably Suvorov!", consider the genus to (Bdn. 6 1-97) were from seed of Algerian ac­ contain a greater number of species. cessions. Seed of M. italica (Bdn. 523) came The chromosome number of many Me li­ from a collection made in Portugal. Seed of Theauthors are. respectively. former postdoctoral lotus species, 2n = 2x = 16, was determined commercially available cultivars of M . alba researchassociate. and professor. Department of Plant ea rly in this century-'U . Karyotype analyses (Evergreen-Lot 14C) and M. officinalis Pathology. Throckmorton Hall. Kansas State Univer­ of all Melilotu s species, with the exception of (Y ukon-F.e. 40594) were used in the study. sity. Munhattun, KS 66506: supervisory research geneticist. USDA-ARS. and George Holmes Professor the du bious M . bicolor Boiss & Bal., were Actively growing root tips from one­ of Agronomy. Departm ent of Agronomy. University reported by Kita5.6. Chromosomes with sat­ mont h-old seedlings grown in clay pots were or Nebraska. Lincoln. NE 68583. Dr. Schlarbaum's ellites were found in all species except M. in­ utilized. The root tips were placed in ice water present address: Depart ment of Forestry. Wildlife and Fisheries. University of Tennessee. Knoxville. TN festa Guss., M. macrocarpa Goss. et Dur., and for 24 hours (for a detailed descr iption, see 37901. Research supported by National Science M . ita lica ( L.) Larn., which belong to Mi­ Singh and Tsuchiyal-) and then fixed in a 3:1 Foundation grant PCM 8022556. Contribu tion no. cromelilotus. Secondary constrictions could mixture of ethanol:propionic acid in which a 82-57 5·). Department of Plant Pathology. Kansas State not be defined in the chromosomes of these small amount of ferric chloride had been dis­ University. Agricultural Experiment Station. Man­ hauan, Kansas 66506. Research was done in coopera­ three species. solved as a mordant. After 3- 4 days in a fix­ tionwiththe Nebraska Agricultural Experiment Sta­ The present study was undert aken to de­ ative, the root tips were hydrolyzed in IN HCI tionand the U.S. Department of Agriculture. Agri­ lineate the occurrence and positions of sec­ at 60°C for 6 minutes and stained in aceto­ cultural Research Service. Please address reprint re­ queststoDr. S. E. Schlar baum. ondary constrictions in the chromosome carmine for at least 3 days. After sufficient i£ 1984. Ameriean Genetic Association. complements of M . infesta, M . macrocarpa , staining, the meristematic tissue was squashed 23 in 45 percent acetic acid. Dividing cells in the meristematic region were isolated and ana­ lyzed. For the Micromelilotu s species, the karyological interpretations were made on the chromosome complement of a representative cell from each species in which the chromo­ somes were well spread and the secondary constrictions were obvious. From these cells, karyotypes were constructed by arranging the I chromosomes in pairs considering the total \ length, centromere position, and gross mor­ phology. The chromosome pairs were gener­ ally arranged in gradation from the longest to A the shortest pair of the complement. Satellite 5p.m chromosome morphology in M. alba and M. officinalis cells was studied in a number of preparations. II 1 11 , I Results 'I'. B The chromosome number of 2n = 2x = 16, as previously rep orted-P-U, was confirmed in FIGU RE 1 A shows somatic chro mosomes of Melilotus infesta, 2n = 2x = 16; all species. Numerous cellswere observed (ca. arrows indicate sate llite chromosomes. B-kar yotype of M . infesta; chromosome s 20) in each species in which chromosome cut from A . morphology was able to be characterized. Secondary constrictions in one chromosome pair were found to exist in M. infe sta , M . macrocarpa, and M. italica (Figures I, 2, 3, yotype were metacentric while the smaller Both species have small satellite bode and 4A -C). The existence of satellite chro­ chromosomes tended to have nonmedian tached to the short ar ms (Figure 4£ E) . mosomes in M. alba 3 and M. offici nalis I also centromeres (Figure 3B) . The chromosomes of M. italica stained very deeply, suggesting was reaffirmed (Figure 4D and E). Discussion The chromosome complement of M. inf esta a heterochromatic natur e. is shown in Figure IA. A secondary constric­ The satellite chromosomes of M. alba and Comparisons of the karyotypes of,\ tion was found in the largest chromosomepair M. officinalis are different in morphology festa , M. macrocarpa, and M. itali: near the centromere in the short arm (Figures from the preceding species (Figure 4A - E ). served in the present study with thos; IB and 4A). At somatic prophase, this chro­ mosome pair appeared to be heterochromatic in contrast to the majority of other chromo­ somes that appeared to stain normally (Figure 5). The karyotypeshows the presence of many nonmetacentric chromosomes (Figure IB). .. Figure 2A showsthe somatic chromosomes of M . macrocarpa. The chromosomes with satellites in this species were the largest chromosomes in the complement (Figure 2B) and appear to be heterochromatic, as in M . •-, infesta. In general, the karyotype of M. ma­ crocarpa closely resembles the M. infesta karyotype. However, theshort arm bearing the satellite body in M. macrocarpa was shorter, A relatively, than the corresponding short arm in M. infesta (Figure 4A and B). The chro­ mosomes of M . macrocarpa, excluding the satellite chromosome pair, did not stain as deeply as chromosomes of the other species. Overall, the chromosomes of M. italica , were the largest of the speciesstudied (Figure 3A). The satellite bodies in this species were B the same or of greater length than the long arms of the satellite chromosomes(Figures 3B FIG URE 2 A shows somatic chromosomes of Melilotus macrocarpa, 2n = 2x = 16; arro» and 4C) . The larger chromosomes in the kar- sate llite chromosomes. B- karyotype of M. macrocarpa; chromosomes cut from A. 24 The Journal of Heredit y Schl arb aum et al.: Seconda ry constrictions in ,~ e4 ) II .... I A •• .. B" .. ' .. I I 5p.m C I'II 'I II B 111111111111 o ,FIGURE 3 A shows somatic chromosomes of Melilotus italica, 2n = 2x = 16;arrows indicate satellite chromosomes. B-karyotype of M. italica; chromosomes cut from A. I E 5,um sen ted by Kita" show a general agreement each species and to artifacts normally en­ FIGURE 4 Satellite chromosomes of Melilotus species isolated from different somatic cells. A--M. with the exception of the satellite chromo­ countered