© Georg Thieme Verlag KG Stuttgart · New York
Development of Simple Sequence Repeat Markers in Cymbopogon Species
Received: December 5, 2005
Accepted: December 22, 2006
22 February 2007 (online)
The genus Cymbopogon comprises about 140 species, which produce characteristic aromatic essential oils. However, the phenotypic identification of species of Cymbopogon has been difficult as a result of widespread occurrence of natural variants, which differ in ploidy levels and chemotaxonomic complexities. Therefore, we have developed a set of simple sequence repeat markers from a genomic library of Cymbopogon jwarancusa to help in the precise identification of the species (including accessions) of Cymbopogon. For this purpose, we isolated 16 simple sequence repeat containing genomic deoxyribonucleic acid clones of C. jwarancusa, which contained a total of 32 simple sequence repeats with a range of 1 to 3 simple sequence repeats per clone. The majority (68.8 %) of the 32 simple sequence repeats comprised dinucleotide repeat motifs followed by simple sequence repeats with trinucleotide (21.8 %) and other higher order repeat motifs. Eighteen (81.8 %) of the 22 designed primers for the above simple sequence repeats amplified products of expected sizes, when tried with genomic DNA of C. jwarancusa, the source species. Thirteen (72.2 %) of the 18 functional primers detected polymorphism among the three species of Cymbopogon (C. flexuosus, C. pendulus and C. jwarancusa) and amplified a total of 95 alleles (range 1 - 18 alleles) with a PIC value of 0.44 to 0.96 per simple sequence repeat. Thus, the higher allelic range and high level of polymorphism demonstrated by the newly developed simple sequence repeat markers are likely to have many applications such as in improvement of essential oil quality by authentication of Cymbopogon species and varieties and mapping or tagging the genes controlling agronomically important traits of essential oils, which can further be utilized in marker assisted breeding.
SSR: Simple sequence repeat
DNA: Deoxyribonucleic acid
PCR: Polymerase chain reaction
dNTP: Deoxynucleotide triphosphate
EST: Express sequence tag
QTL: Quantitative trait loci
PIC: Polymorphic information content
MAB: Marker assisted selection
Poaceae - Cymbopogon species - genomic library - SSR markers - polymorphism
- 1 Khanuja S PS, Shasany A K, Pawar A, Lal R K, Darokar M P, Naqvi A A. et al . Essential oil constituents and RAPD markers to establish species relationships in Cymbopogon Spreng. (Poaceae). Biochem Syst Ecol. 2005; 33 171-86.
- 2 Joshi K, Chavan P, Warude D, Patwardhan B. Molecular markers in herbal drug technology. Curr Sci. 2004; 87 159-65.
- 3 Rao B L. Scope for development of new cultivars of Cymbopogons as a source of terpene chemicals. In: Handa, SS, Kaul MK, editors
Supplement to cultivation and utilization of aromatic plants.New Delhi NISC; Dr. K.S. Krishnan Marg 1997: 71-83.
- 4 Yang M, Zhang D, Liu J, Zheng J. A molecular marker that is specific to medicinal rhubarb based on chloroplast trnL/trnF sequences. Planta Med. 2001; 67 784-6.
- 5 Li Y F, Li Y X, Lin J, Xu Y, Yan F, Tang L. et al . Identification of bulb from Fritillaria cirrhosa by PCR with specific primers. Planta Med. 2003; 69 186-8.
- 6 Crockett S L, Douglas A W, Scheffler B E, Khan I A. Genetic profiling of Hypericum (St. John’s Wort) species by nuclear ribosomal ITS sequence analysis. Planta Med. 2004; 70 929-35.
- 7 Gupta P K, Balyan H S, Sharma P C, Ramesh B. Microsatellite in plants: a new class of molecular markers. Curr Sci. 1996; 70 45-54.
- 8 Cordeiro G M, Taylor G O, Henry R J. Characterization of microsatellite markers from sugarcane (Saccharum sp.), a highly polyploid species. Plant Sci. 2000; 155 161-8.
- 9 Schlotterer C, Tautz D. Slippage synthesis of simple sequence DNA. Nucleic Acids Res. 1992; 20 211-5.
- 10 Prabhu K V, Somers D J, Rakow G, Gugel R K. Molecular markers linked to white rust resistance in mustard Brassica juncea . Theor Appl Genet. 1998; 97 865-70.
- 11 Zane L, Bargelloni L, Patarnello T. Strategies for microsatellite isolation: a review. Mol Ecol. 2002; 11 1-6.
- 12 Tegelstrom H. Detection of mitochondrial DNA fragments. In: Hoelzel AR, editor. Molecular genetic analysis of populations: a practical approach. Oxford, New York: IRL. Press; 1992 89-114.
- 13 Weir B S. Genetic-data analysis methods for discrete genetic data. Sunderland; Sinauer Association Inc 1990.
- 14 Ashkenazi V, Chani E, Lavi U, Levy D, Hillel J, Veilleux R E. Development of microsatellite markers in potato and their use in phylogenetics and fingerprinting analyses. Genome. 2001; 44 50-62.
- 15 Brown S M, Hopkins M S, Mitchell S E, Senior M L, Wang T Y, Duncan R R. et al . Multiple method for the identification of polymorphic simple sequence repeats (SSRs) in sorghum [Sorghum bicolor (L.) Moench]. Theor Appl Genet. 1996; 93 190-8.
- 16 Cordeiro G M, Cazu R, Me Intigre C L, Manners J M, Henry R J. Microsatellite markers for sugarcane (Saccharum spp.) ESTs cross transferable to erianthus an sorghum. Plant Sci. 2001; 160 1115-23.
- 17 Stajner N, Jakse J, Kozjak P, Javornik B. The isolation and characterization of microsatellites hop (Humulus lupulus L.) Plant Sci. 2004; 168 213-21.
- 18 Powell W, Machrary G C, Rovan J. Polymorphism revealed by simple sequence repeats. Trends Plant Sci. 1996; 1 215-22.
Dr. Jitendra Kumar
Department of Genetics & Plant Breeding
Ch. Charan Singh University