Unlocking Genetic Diversity and Germplasm Characterization with Molecular Markers: Strategies for Crop Improvement

Aniket Bunjkar *

Department of Genetics and Plant Breeding, Lovely Professional University, Phagwara, Punjab, 144411, India.

Puneet Walia

Department of Genetics and Plant Breeding, Lovely Professional University, Phagwara, Punjab, 144411, India.

Sanjeet Singh Sandal

Department of Genetics and Plant Breeding, Lovely Professional University, Phagwara, Punjab, 144411, India.

*Author to whom correspondence should be addressed.


Molecular markers have emerged as the most useful tools in assessment of genetic diversity and characterizing germplasm, for crop improvement. This review paper comprehensively analyzes the applications of molecular markers in diversity analysis and germplasm characterization. It underscores the significance of genetic diversity as the bedrock for plant breeding programs, enabling the development of improved varieties with desirable attributes such as higher yields, stress tolerance, and enhanced nutritional profiles. The paper provides an overview of various types of molecular markers, including hybridization-based markers (e.g., RFLPs) and PCR-based markers (e.g., RAPDs, AFLPs, SSRs, and SNPs). It discusses marker selection strategies, emphasizing the consideration of factors like polymorphism, informativeness, and the potential for multiplexing and high-throughput genotyping. Diversity analysis techniques, including principal component analysis (PCA), cluster analysis methods (UPGMA and Neighbor-Joining), and population structure analysis (model-based approaches like STRUCTURE), are detailed. These methods enable the assessment of genetic relationships, identification of subpopulations, and selection of diverse parents based on molecular marker data. The review further explores the applications of molecular markers in germplasm management, conservation, and utilization. It discusses the role of markers in targeted introgression of desirable traits from diverse sources, as well as the integration of genotypic and phenotypic data for association mapping, genomic prediction, and selection. Additionally, the paper addresses emerging technologies, such as next-generation sequencing (NGS) and the integration of molecular markers with other omics data. It also highlights the practical applications and impact of molecular markers in crop improvement programs, including marker-assisted selection (MAS), genomic selection, and the development of genetically modified crops. Finally, the review outlines challenges and future perspectives, including limitations0 of current technologies, the potential of emerging techniques like NGS, and the integration of molecular markers with other omics approaches for a comprehensive understanding of complex traits.

Keywords: Molecular markers, genetic diversity, germplasm characterization, principal component analysis, cluster analysis, population structure

How to Cite

Bunjkar , A., Walia , P., & Sandal , S. S. (2024). Unlocking Genetic Diversity and Germplasm Characterization with Molecular Markers: Strategies for Crop Improvement. Journal of Advances in Biology & Biotechnology, 27(6), 160–173. https://doi.org/10.9734/jabb/2024/v27i6873


Download data is not yet available.


Cole CT. Genetic variation in rare and common plants. Annual Review of Ecology, Evolution, and Systematics. 2009;34(1):213-237.

Nadeem MA, Nawaz MA, Shahid MQ, Doğan Y, Comertpay G, Yıldız M, Baloch FS. DNA molecular markers in plant breeding: current status and recent advancements in genomic selection and genome editing. Biotechnology & Biotechnological Equipment. 2018;32(2): 261-285.

Zebire DA. Applications of molecular markers in Genetic Diversity Studies of maize. Nigerian Journal of Biotechnology. 2020;37(1):101-108.

Brown WL. Genetic diversity and genetic vulnerability—An appraisal. Economic botany. 1983;37:4-12.

Ennos R. Genetic variation and conservation of British native trees and shrubs: Current knowledge and policy implications; 2000.

Swingland IR. Biodiversity, definition of. Encyclopedia of biodiversity. 2001;1:377-391.

Frankham R, Briscoe DA, Ballou JD. Introduction to conservation genetics. Cambridge University Press; 2002.

Hughes AR, Inouye BD, Johnson MT, Underwood N, Vellend M. Ecological consequences of genetic diversity. Ecology letters. 2008;11(6):609-623.

Swarup S, Cargill EJ, Crosby K, Flagel L, Kniskern J, Glenn KC. Genetic diversity is indispensable for plant breeding to improve crops. Crop Science. 2021;61(2):839-852.

Bhandari HR, Bhanu AN, Srivastava K, Singh MN, Shreya HA. Assessment of genetic diversity in crop plants-an overview. Adv. Plants Agric. Res. 2017; 7(3):279-286.

Raza A, Razzaq A, Mehmood SS, Zou X, Zhang X, Lv Y, Xu J. Impact of climate change on crops adaptation and strategies to tackle its outcome: A review. Plants. 2019;8(2):34.

Raffard A, Santoul F, Cucherousset J, Blanchet S. The community and ecosystem consequences of intraspecific diversity: A meta‐analysis. Biological Reviews. 2019;94(2):648-661.

Ramanatha Rao V, Hodgkin T. Genetic diversity and conservation and utilization of plant genetic resources. Plant cell, tissue and organ culture. 2002;68:1-19.

Dwivedi SL, Ceccarelli S, Blair MW, Upadhyaya HD. Are AK, Ortiz R. Landrace germplasm for improving yield and abiotic stress adaptation. Trends in plant science. 2016;21(1):31-42.

Food and agriculture organization of the United Nations. Genebank Standards for Plant Genetic Resources for Food and Agriculture; 2014. Retrieved from: https://www.fao.org/3/ i3704e/i3704e.pdf

Begna T, Begna T. Role and economic importance of crop genetic diversity in food security. International Journal of Agricultural Science and Food Technology. 2021;7(1):164-169.

Salgotra RK, Chauhan BS. Genetic diversity, conservation, and utilization of plant genetic resources. Genes. 2023; 14(1):174.

Saad NSM, Neik TX, Thomas WJ, Amas JC, Cantila AY, Craig RJ, Batley J. Advancing designer crops for climate resilience through an integrated genomics approach. Current Opinion in Plant Biology. 2022;67:102220.

Mondal R, Kumar A, Gnanesh BN. Crop germplasm: Current challenges, physiological-molecular perspective, and advance strategies towards development of climate-resilient crops. Heliyon. 2023; 9(1).

Mondini L. Noorani A, Pagnotta MA. Assessing plant genetic diversity by molecular tools. Diversity. 2009;1(1):9-35.

Mirzaei S. Application of molecular markers in plant sciences; An overview. Central Asian Journal of Plant Science Innovation. 2021;1(4):192-200.

Amom T, Nongdam P. The use of molecular marker methods in plants: a review. International Journal of Current Research and Review. 2017;9(17):1-7.

Dhutmal RR, Mundhe AG, More AW. Molecular marker techniques: A Review. Int. J. Curr. Microbiol. Appl. Sci. 2018;6: 816-825.

Botstein D, White RL, Skolnick M, Davis RW. Construction of a genetic linkage map in man using restriction fragment length polymorphisms. American journal of human genetics. 1980;32(3):314.

Jiang GL. Molecular markers and marker-assisted breeding in plants. Plant breeding from laboratories to fields. 2013;3:45-83.

Singh BD, Singh AK. Marker-assisted plant breeding: principles and practices; 2015.

Williams JG, Kubelik AR, Livak KJ, Rafalski JA, Tingey SV. DNA polymorphisms amplified by arbitrary primers are useful as genetic markers. Nucleic acids research. 1990;18(22), 6531-6535.

Welsh J, McClelland M. Fingerprinting genomes using PCR with arbitrary primers. Nucleic acids research. 1990;18(24):7213-7218.

Babu KN, Sheeja TE, Minoo D, Rajesh MK, Samsudeen K, Suraby EJ, Kumar IPV. Random amplified polymorphic DNA (RAPD) and derived techniques. Molecular Plant Taxonomy: Methods and Protocols. 2021;219-247.

Jones CJ, Edwards KJ, Castaglione S, Winfield MO, Sala F. Van de Wiel C, Karp A. Reproducibility testing of RAPD, AFLP and SSR markers in plants by a network of European laboratories. Molecular breeding. 1997;3:381-390.

Lynch M, Walsh B. Genetics and analysis of quantitative traits. Sunderland, MA: Sinauer. 1998;1:535-557.

Vos P, Hogers R, Bleeker M, Reijans M, Lee TVD, Hornes M, Zabeau M. AFLP: A new technique for DNA fingerprinting. Nucleic acids research. 1995;23(21):4407-4414.

Madhumati B. Potential and application of molecular markers techniques for plant genome analysis. Int J Pure App Biosci. 2014;2(1):169-188.

Litt M, Luty JA. A hypervariable microsatellite revealed by in vitro amplification of a dinucleotide repeat within the cardiac muscle actin gene. American journal of human genetics. 1989;44(3):397.

Tautz D. Hypervariability of simple sequences as a general source for polymorphic DNA markers. Nucleic acids research. 1989;17(16):6463-6471.

Tautz D, Ren M. Simple sequences are ubiquitous repetitive components of eukaryotic genomes. Nucleic acids research. 1984;12(10):4127-4138.

Mir RR, Varshney RK. 10 Future prospects of molecular markers in Plants. Molecular markers in plants. 2013;169.

Guichoux E, Lagache L, Wagner S, Chaumeil P, Léger P, Lepais O, Petit RJ. Current trends in microsatellite genotyping. Molecular ecology resources. 2011;11(4): 591-611.

Xu Y. Molecular plant breeding. Cabi; 2010.

Sobrino B, Brión M, Carracedo A. SNPs in forensic genetics: A review on SNP typing methodologies. Forensic science international. 2005;154(2-3):181-194.

Hildebrand CE, Torney DC, Wagner RP. Informativeness of polymorphic DNA markers. Los Alamos Science. 1992; 20(20):100-102.

Vieira MLC, Santini L, Diniz AL, Munhoz CDF. Microsatellite markers: what they mean and why they are so useful. Genetics and molecular biology. 2009; 32(3):308-328.

Rasheed A, Hao Y, Xia X, Khan A, Xu Y, Varshney RK, He Z. Crop breeding chips and genotyping platforms: progress, challenges, and perspectives. Molecular plant. 2017;10(8):1047-1064.

Mohammadi SA, Prasanna BM. Analysis of genetic diversity in crop plants—salient statistical tools and considerations. Crop science. 2003;43(4):1235-1248.

Nei M. Analysis of gene diversity in subdivided populations. Proceedings of the national academy of sciences. 1973; 70(12):3321-3323.

Mammadov J. Aggarwal R, Buyyarapu R, Kumpatla S. SNP markers and their impact on plant breeding. International journal of plant genomics; 2012.

Thachuk C, Crossa J, Franco J, Dreisigacker S, Warburton M, Davenport GF. Core hunter: an algorithm for sampling genetic resources based on multiple genetic measures. BMC bioinformatics. 2009;10(1):1-12.

Semagn K, Babu R, Hearne S, Olsen M. Single nucleotide polymorphism genotyping using Kompetitive Allele Specific PCR (KASP): overview of the technology and its application in crop improvement. Molecular Breeding. 2014; 33(1):1-14.

Hayden MJ, Nguyen TM, Waterman A, Chalmers KJ. Multiplex-ready PCR: a new method for multiplexed SSR and SNP genotyping. BMC genomics. 2008;9(1):1-12.

Schuelke M. An economic method for the fluorescent labeling of PCR fragments. Nature Biotechnology. 2000;18(2):233-234.

Henegariu O, Heeresen NA. Dlouhy SR. Vance GH, Vogt PH. Multiplex PCR: Critical parameters and step-by-step protocol. Biotechniques. 1997;23(3):504-511.

Wenzl P, Carling J, Kudrna D, Jaccoud D, Huttner E, Kleinhofs A, Kilian A. Diversity Arrays Technology (DArT) for whole-genome profiling of barley. Proceedings of the National Academy of Sciences. 2004; 101(26):9915-9920.

Davey JW, Hohenlohe PA, Etter PD, Boone JQ, Catchen JM, Blaxter ML. Genome-wide genetic marker discovery and genotyping using next-generation sequencing. Nature Reviews Genetics. 2011;12(7):499-510.

Huang X, Wei X, Sang T, Zhao Q, Feng Q, Zhao Y, Han B. Genome-wide association studies of 14 agronomic traits in rice landraces. Nature Genetics. 2010;42(11): 961-967.

Crossa J, Pérez-Rodríguez P, Cuevas J, Montesinos-López O, Jarquín D, de los Campos G, Burgueño J. Genomic selection in plant breeding: Methods, models, and perspectives. Crop Science. 2017;57(3):1058-1067.

Huang, X., Han, B., & Sunderman, D. Analysis of genetic diversity in sweet potato (Ipomoea batatas) germplasm resources explored by genomic markers. Genetics and Molecular Biology. 2015; 34(4)590-596.

Dufresne F, Stift M, Vergilino R, Mable BK. Recent progress and challenges in population genetics of polyploid organisms: an overview of current state-of-the-art molecular and statistical tools. Molecular Ecology. 2019;23(1):40-69.

Jombart T, Pontier D, Dufour AB. Genetic markers in the playground of multivariate analysis. Heredity. 2009;102(4):330-341.

Meirmans PG. Seven common mistakes in population genetics and how to avoid them. Molecular Ecology. 2015;24(13): 3223-3231.

Price AL, Patterson NJ, Plenge RM, Weinblatt ME, Shadick NA, Reich D. Principal components analysis corrects for stratification in genome-wide association studies. Nature Genetics. 2006;38(8):904-909.

Zou C, Fu Y, Song W, Mei X, Liu J, Tang H. Principal component analysis combined with a model-based approach dissects genetic structure of germplasm populations: A case study of rice. Crop Science. 2021;61(5):3030-3041.

Chakravarthi BK, Naravaneni R. SSR marker based DNA fingerprinting and diversity study in rice (Oryza sativa. L). African Journal of Biotechnology. 2006; 5(9):684-688.

Malosetti M, Ribaut JM, van Eeuwijk FA. The statistical analysis of multi-environment data: Modeling genotype-by-environment interaction and its genetic basis. Frontiers in physiology. 2007;4:44.

Sneath PHA. The principles and practice of numerical classification. Numerical taxonomy. 1973;573.

Saitou N, Nei M. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution. 1987;4(4):406-425.

Takezaki N, Nei M. Empirical tests of the reliability of phylogenetic trees constructed with microsatellite DNA. Genetics, 2008; 178(1):385-392.

Ghasemi AR, Golparvar AR, Isfahani MN. Analysis of genetic diversity of sugar beet genotypes using random amplified polymorphic DNA marker. Genetika. 2014; 46(3):975-984.

Liu K, Muse SV. PowerMarker: an integrated analysis environment for genetic marker analysis. Bioinformatics. 2005; 21(9):2128-2129.

Perrier X. DARwin software; 2006. Available:http://darwin.cirad.fr/darwin.

Kumar H, Kaur G, Banga S. Molecular characterization and assessment of genetic diversity in sesame (Sesamum indicum L.) germplasm collection using ISSR markers. Journal of crop improvement. 2012 Jul 1;26(4):540-57.

Pritchard JK, Stephens M, Donnelly P. Inference of population structure using multilocus genotype data. Genetics. 2000; 155(2):945-959.

Raj A, Stephens M, Pritchard JK. Fast structure: Variational inference of population structure in large SNP data sets. Genetics. 2014;197(2):573-589.

Corander J, Waldmann P, Marttinen P, Sillanpää MJ. BAPS 2: Enhanced possibilities for the analysis of genetic population structure. Bioinformatics. 2004; 20(15):2363-2369.

Pella J, Masuda M. The gibbs and split merge sampler for population mixture analysis from genetic data with incomplete baselines. Canadian Journal of Fisheries and Aquatic Sciences. 2006;63(3):576-596.

François O, Ancelet S, Guillot G. Bayesian clustering using hidden Markov random fields in spatial population genetics. Genetics. 2006;174(2):805-816.

Guillot G, Santos F, Estoup A. Analysing georeferenced population genetics data with Geneland: a new algorithm to deal with null alleles and a friendly graphical user interface. Bioinformatics. 2008;24(11): 1406-1407.

Gao H, Williamson S, Bustamante CD. A markov chain monte carlo approach for joint inference of population structure and inbreeding rates from multilocus genotype data. Genetics. 2007;176(3):1635-1651.

Ma M, Lei E, Meng H, Wang T, Xie L, Shen D, Lu B. Cluster and principal component analysis based on SSR markers of Amomum tsao-ko in Jinping County of Yunnan Province. In AIP Conference Proceedings. (Vol. 1864, No. 1). AIP Publishing; 2017.

Hammer O. PAST: Paleontological statistics software package for education and data analysis. Palaeontol Electron. 2001;4:9.

Patwardhan A, Ray S, Roy A. Molecular markers in phylogenetic studies-a review. Journal of Phylogenetics & Evolutionary Biology. 2014;2(2):131.

Odong TL, Jansen J, Van Eeuwijk FA, Van Hintum TJ. Quality of core collections for effective utilisation of genetic resources review, outlook and guidelines. Biotechnology and Genetic Engineering Reviews. 2013;29(1):135-156.

Govindaraj M, Vetriventhan M, Srinivasan M. Importance of genetic diversity assessment in crop plants and its recent advances: An overview of its analytical perspectives. Genetics Research International. 2015;1-14.

Liu X, Liu H, Ma L, Li X, Xu C, Su H, Fan Y. Construction of sugarcane hybrids core collection by using stepwise clustering sampling approach with molecular marker data. Acta Agronomica Sinica. 2014; 40(11):1885-1894.

Pandey MK, Roorkiwal M, Singh VK, Ramalingam A, Kudapa H, Thudi M, Varshney RK. Emerging genomic tools for legume breeding: current status and future prospects. Frontiers in Plant Science. 2016;7:455.

Collard BCY, Mackill DJ. Marker-assisted selection: an approach for precision plant breeding in the twenty-first century. Philosophical Transactions of the Royal Society B: Biological Sciences. 2008; 363(1491):557-572.

Xu Y, Crouch JH. Marker-assisted selection in plant breeding: from publications to practice. Crop Science. 2008;48(2):391-407.

Langridge P, Waugh R. Harnessing the potential of germplasm collections. Nature Genetics. 2019;51(2):200-201.

Xu Y, Li P, Yang Z, Xu C. Genetic mapping of quantitative trait loci in crops. The Crop Journal. 2017;5(2):175-184.

Bernardo R. Molecular markers and selection for complex traits in plants: Learning from the last 20 years. Crop Science. 2016;56(1):53-65.

Kumawat G, Singh G, Gireesh C, Shivakumar M, Arya M, Agarwal DK, Husain SM. Molecular characterization and genetic diversity analysis of soybean (Glycine max (L.) Merr.) germplasm accessions in India. Physiology and molecular biology of plants. 2015;21:101-7.

Korte A, Farlow A. The advantages and limitations of trait analysis with GWAS: A review. Plant Methods. 2013;9(1):1-9.

Huang X, Han B. Natural variations and genome-wide association studies in crop plants. Annual review of plant biology. 2014;65:531-551.

Juliana P, Singh RP, Singh PK, Crossa J, Huerta-Espino J, Lan C, Sorrells ME. Genomic and pedigree-based prediction for leaf, stem, and stripe rust resistance in wheat. Theoretical and Applied Genetics. 2018;131(8):1691-1701.

de los Campos G, Hickey JM, Pong-Wong R, Daetwyler HD, Calus MP. Whole-genome regression and prediction methods applied to plant and animal breeding. Genetics. 2013;193(2):327-345.

Meuwissen TH, Hayes BJ, Goddard ME. Prediction of total genetic value using genome-wide dense marker maps. Genetics. 2001;157(4):1819-1829.

Hickey JM, Chiurugwi T, Mackay I, Powell W, Eggen A, Kilian A, Gorjanc G. Genomic prediction unifies animal and plant breeding programs to form platforms for biological discovery. Nature Genetics. 2017;49(9):1297-1303.

Deschamps S, Llaca V, May GD. Genotyping-by-sequencing in plants. Biology. 2012;1(3):460-483.

He J, Zhao X, Laroche A, Lu ZX, Liu H, Li Z. Genotyping-by-sequencing BS), an ultimate marker-assisted selection (MAS) tool to accelerate plant breeding. Frontiers in Plant Science. 2014;5:484.

Huang X, Feng Q, Qian Q, Zhao Q, Wang L, Wang A, Han B. High-throughput genotyping by whole-genome resequencing. Genome Research. 2014; 24(7):1183-1183.

Varshney RK, Graner A, Sorrells ME. Genic microsatellite markers in plants: Features and applications. Trends in Biotechnology. 2009;23(1):48-55.

Langridge P, Waugh R. Harnessing the potential of germplasm collections. Nature Genetics. 2019;51(2):200-201.

Tester M, Langridge P. Breeding technologies to increase crop production in a changing world. Science. 2010; 327(5967):818-822.

Abberton M, Batley J, Bentley A, Bryant J, Cai H, Cairns J, Edwards, D. Global agricultural intensification during climate change: a role for genomics. Plant Biotechnology Journal. 2016;14(4):1095-1098.