Skip to main content
SpringerLink
Log in

Cloning and characterization of chromosomal markers in alfalfa (Medicago sativa L.)

  • Original Paper
  • Published:
Theoretical and Applied Genetics Aims and scope Submit manuscript

Abstract

Eleven tandemly repetitive sequences were identified from a Cot-1 library by FISH and sequence analysis of alfalfa (Medicago sativa). Five repetitive sequences (MsCR-1, MsCR-2, MsCR-3, MsCR-4, and MsCR-5) were centromeric or pericentromeric, of which three were satellite DNAs and two were minisatellite DNAs. Monomers of 144, 148, and 168 bp were identified in MsCR-1, MsCR-2, and MsCR-3, respectively, while 15 and 39 bp monomers were identified in MsCR-4 and MsCR-5, respectively. Three repetitive sequences were characterized as subtelomeric; one repetitive sequence, MsTR-1, had a 184 bp monomer, and two repetitive sequences had fragments of 204 and 327 bp. Sequence analysis revealed homology (70–80 %) between MsTR-1 and a highly repeated sequence (C300) isolated from M. ssp. caerulea. Three identified repetitive sequences produced hybridization signals at multiple sites in a few of the chromosomes; one repetitive sequence was identified as the E180 satellite DNA previously isolated from M. sativa, while the other 163 and 227 bp fragments had distinct sequences. Physical mapping of the repetitive sequences with double-target FISH revealed different patterns. Thus, nine novel tandemly repetitive sequences that can be adopted as distinct chromosome markers in alfalfa were identified in this study. Furthermore, the chromosome distribution of each sequence was well described. Though significant chromosome variations were detected within and between cultivars, a molecular karyotype of alfalfa was suggested with the chromosome markers we identified. Therefore, these novel chromosome markers will still be a powerful tool for genome composition analysis, phylogenetic studies, and breeding applications.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  • Allen G, Flores-Vergara M, Krasynanski S, Kumar S, Thompson W (2006) A modified protocol for rapid DNA isolation from plant tissues using cetyltrimethylammonium bromide. Nat Protoc 1:2320–2325

    Article  PubMed  CAS  Google Scholar 

  • Barnes D, Bingham E, Murphy R, Hunt O, Beard D, Skrdla W, Teuber L (1977) Alfalfa germplasm in the United States: genetic vulnerability, use, improvement and maintenance. USDA Technical Bulletin 1571. ARS, Washington DC, p 21

    Google Scholar 

  • Bauchan G, Hossain M (1997) Identification of Alfalfa chromosomes using Geimsa banding and image analysis techniques. In: Proceedings of the XVIII International Rangeland Congress, Canada, pp 61–62

  • Bauchan GR, Hossain MA (2001) Distribution and characterization of heterochromatic DNA in the tetraploid African population alfalfa genome. Crop Sci 41:1921–1926

    Article  Google Scholar 

  • Begum R, Alam SS, Menzel G, Schmidt T (2009) Comparative molecular cytogenetics of major repetitive sequence families of three Dendrobium species (Orchidaceae) from Bangladesh. Ann Bot 104:863–872

    Article  PubMed  CAS  Google Scholar 

  • Blondon F, Marie D, Brown S, Kondorosi A (1994) Genome size and base composition in Medicago sativa and Medicago truncatula species. Genome 37:264–270

    Article  PubMed  CAS  Google Scholar 

  • Calderini O, Pupilli F, Paolocci F, Arcioni S (1997) A repetitive and species-specific sequence as a tool for detecting the genome contribution in somatic hybrids of the genus Medicago. Theor Appl Genet 95:734–740

    Article  CAS  Google Scholar 

  • Cheng Z, Dong F, Langdon T, Ouyang S, Buell CR, Gu M, Blattner FR, Jiang J (2002) Functional rice centromeres are marked by a satellite repeat and a centromere-specific retrotransposon. Plant Cell 14:1691–1704

    Article  PubMed  CAS  Google Scholar 

  • Cleveland R, Stanford E (1959) Chromosome pairing in hybrids between tetraploid Medicago sativa L. and diploid Medicago falcata L. Agron J 51:488–492

    Article  Google Scholar 

  • Dou QW, Chen ZG, Liu YA, Tsujimoto H (2009) High frequency of karyotype variation revealed by sequential FISH and GISH in plateau perennial grass forage Elymus nutans. Breed Sci 59:651–656

    Article  CAS  Google Scholar 

  • Falistocco E (2000) Brief communication. Physical mapping of rRNA genes in Medicago sativa and M. glomerata by fluorescent in situ hybridization. J Hered 91:256–260

    Article  PubMed  CAS  Google Scholar 

  • Falistocco E, Falcinelli M, Veronesi F (1995) Karyotype and C-banding pattern of mitotic chromosomes in alfalfa, Medicago sativa L. Plant Breed 114:451–453

    Article  Google Scholar 

  • Fonsêca A, Ferreira J, dos Santos TR, Mosiolek M, Bellucci E, Kami J, Gepts P, Geffroy V, Schweizer D, dos Santos KGB, Pedrosa-Harand A (2010) Cytogenetic map of common bean (Phaseolus vulgaris L.). Chromosome Res 18:487–502

    Article  PubMed  Google Scholar 

  • Fukui K, Kamisugi Y, Sakai F (1994) Physical mapping of 5S rDNA loci by direct-cloned biotinylated probes in barley chromosomes. Genome 37:105–111

    Article  PubMed  CAS  Google Scholar 

  • Galasso I, Schmidt T, Pignone D (2001) Identification of Lens culinaris ssp. culinaris chromosomes by physical mapping of repetitive DNA sequences. Chromosome Res 9(3):199–209

    Article  PubMed  CAS  Google Scholar 

  • Gillies CB (1970) Alfalfa Chromosomes 1. Pachytene karyotype of diploid Medicago falcata L. and its relationship to Medicago sativa L. Crop Sci 10:169–171

    Article  Google Scholar 

  • Han Y, Zhang Z, Huang S, Jin W (2011) An integrated molecular cytogenetic map of Cucumis sativus L. chromosome 2. BMC Genet 12:18

    Article  PubMed  Google Scholar 

  • Ho KM, Kasha KJ (1972) Chromosome homology at pachytene in diploid Medicago sativa, Medicago falcata and their hybrids. Can J Genet Cytol 14:829–838

    Google Scholar 

  • Jiang JM, Gill BS (2006) Current status and the future of fluorescence in situ hybridization (FISH) in plant genome research. Genome 49:1057–1068

    Article  PubMed  CAS  Google Scholar 

  • Jin W, Melo JR, Nagaki K, Talbert PB, Henikoff S, Dawe RK, Jiang J (2004) Maize centromeres: organization and functional adaptation in the genetic background of oat. Plant Cell 16:571–581

    Article  PubMed  CAS  Google Scholar 

  • Kato A, Lamb JC, Birchler JA (2004) Chromosome painting using repetitive DNA sequences as probes for somatic chromosome identification in maize. Proc Natl Acad Sci USA 101:13554

    Article  PubMed  CAS  Google Scholar 

  • Kulikova O, Geurts R, Lamine M, Kim DJ, Cook DR, Leunissen J, De Jong H, Roe BA, Bisseling T (2004) Satellite repeats in the functional centromere and pericentromeric heterochromatin of Medicago truncatula. Chromosoma 113:276–283

    Article  PubMed  CAS  Google Scholar 

  • Lesins K (1957) Cytogenetic study on a tetraploid plant at the diploid chromosome level. Can J Bot 35:181–196

    Article  Google Scholar 

  • Masoud SA, Gill BS, Johnson LB (1991) C-Banding of alfalfa chromosomes standard karyotype and analysis of a somaclonal variant. J Hered 82:335–338

    Google Scholar 

  • McCoy TJ, Bingham ET (1988) Cytology and cytogenetics of alfalfa. In: alfalfa and alfalfa improvement. Agron. Monogr. 29. ASA, CSSA and SSSA. Madison, WI, pp 737–776

  • Paesold S, Borchardt D, Schmidt T, Dechyeva D (2012) A sugar beet (Beta vulgaris L.) reference FISH karyotype for chromosome and chromosome-arm identification, integration of genetic linkage groups and analysis of major repeat family distribution. Plant J 72:600–611

    Article  PubMed  CAS  Google Scholar 

  • Richards EJ, Ausubel FM (1988) Isolation of a higher eukaryotic telomere from Arabidopsis thaliana. Cell 53:127–136

    Article  PubMed  CAS  Google Scholar 

  • Rosato M, Galian JA, Rossello JA (2012) Amplification, contraction and genomic spread of a satellite DNA family (E180) in Medicago (Fabaceae) and allied genera. Ann Bot 109:773–782

    Article  PubMed  CAS  Google Scholar 

  • Schmidt T, Heslop-Harrison JS (1998) Genomes, genes and junk: a model of the large-scale organization of plant chromosomes. Trends Plant Sci 3:195–199

    Article  Google Scholar 

  • Winicov I, Maki DH, Waterborg JH, Riehm MR, Harrington RE (1988) Characterization of the alfalfa (Medicago sativa) genome by DNA reassociation. Plant Mol Biol 10:369–371

    Article  CAS  Google Scholar 

  • Xia X, Erickson L (1993) An AT-rich satellite DNA sequence, E180, in alfalfa (Medicago sativa). Genome 36:427–432

    Article  PubMed  CAS  Google Scholar 

  • Zakrzewski F, Wenke T, Holtgrawe D, Weisshaar B, Schmidt T (2010) Analysis of a Cot-1 library enables the targeted identification of minisatellite and satellite families in Beta vulgaris. BMC Plant Bio 10:8

    Article  Google Scholar 

  • Zhang L, Xu C, Yu W (2012) Cloning and characterization of chromosomal markers from a Cot-1 library of peanut (Arachis hypogaea L.). Cytogenet Genome Res 137:31–41

    Article  PubMed  CAS  Google Scholar 

  • Zwick MS, Hanson RE, Islam-Faridi MN, Stelly DM, Wing RA, Price HJ, McKnight TD (1997) A rapid procedure for the isolation of Cot-1 DNA from plants. Genome 40:138–142

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We thank Professor Tao Wang (China Agriculture University) for providing the seeds of some alfalfa materials. This study was financially supported by the “Joint Scholars” program of the “Lights in the Western Region” talent cultivation plan of the Chinese Academy of Sciences and partly supported by the Main Direction Program for Knowledge Innovation of the Chinese Academy of Sciences (KSCX2-EW-Q-23).

Author information

Authors and Affiliations

  1. Key Laboratory of Adaptation and Evolution of Plateau Biota, Northwest Plateau Institute of Biology, Chinese Academy of Sciences, Xining, 810001, China

    Feng Yu, Yunting Lei, Yuan Li, Quanwen Dou, Haiqing Wang & Zhiguo Chen

  2. Graduate University of Chinese Academy of Sciences, Beijing, 100049, China

    Feng Yu & Yuan Li

Authors
  1. Feng Yu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yunting Lei
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Yuan Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Quanwen Dou
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Haiqing Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Zhiguo Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Quanwen Dou.

Additional information

Communicated by B. Friebe.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOC 119 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Yu, F., Lei, Y., Li, Y. et al. Cloning and characterization of chromosomal markers in alfalfa (Medicago sativa L.). Theor Appl Genet 126, 1885–1896 (2013). https://doi.org/10.1007/s00122-013-2103-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00122-013-2103-z

Keywords

Search

Navigation