Abstract
To unravel the plastid genome diversity among the cultivated groups of the pigeonpea germplasm, we characterized the SNP occurrence and distribution of 142 pigeonpea mini-core collections based on their reference-based assembly of the chloroplast genome. A total of 8921 SNPs were found, which were again filtered and finally 3871 non-synonymous SNPs were detected and used for diversity estimates. These 3871 SNPs were classified into 12 groups and were present in only 44 of the 125 genes, demonstrating the presence of a precise mechanism for maintaining the whole chloroplast genome throughout evolution. The Acetyl-CoA carboxylase D gene possesses the maximum number of SNPs (12.29%), but the Adenosine Tri-Phosphate synthatase cluster genes (atpA, atpB, atpE, atpF, atpH, and atpI) altogether bear 43.34% of the SNPs making them most diverse. Various diversity estimates, such as the number of effective alleles (1.013), Watterson’s estimate (0.19), Tajima’s D ( – 3.15), Shannon’s information index (0.036), suggest the presence of less diversity in the cultivated gene pool of chloroplast genomes. The genetic relatedness estimates based on pairwise correlations were also in congruence with these diversity descriptors and indicate the prevalence of rare alleles in the accessions. Interestingly, no stratification was observed either through STRUCTURE, PCoA, or phylogenetic analysis, indicating the common origin of the chloroplast in all the accessions used, irrespective of their geographical distribution. Further 6194 Cleaved Amplified Polymorphic Sequences (CAPS) markers for 531 SNPs were developed and validated in a selected set of germplasm. Based on these results, we inferred that all of the cultivated gene pools of pigeonpea have a common origin for the chloroplast genome and they possess less diversity in protein-coding regions, indicating a stable and evolved plastid genome. At the same time, all diversity analysis indicates the occurrence of rare alleles, suggesting the suitability of the mini-core collection in future pigeonpea improvement programs. In addition, the development of chloroplast genome-based CAPS markers would have utility in pigeonpea breeding programs.









Similar content being viewed by others
Explore related subjects
Discover the latest articles and news from researchers in related subjects, suggested using machine learning.Data availability
All online data are cited and other data related to the manuscript are provided in the manuscript.
References
Abbas B, Renwarin Y, Bintoro A et al (2010) Genetic diversity of sago palm in Indonesia based on chloroplast DNA (cpDNA) markers. Biodivers J Biol Divers https://doi.org/10.13057/biodiv/d110302
Ackerman MS, Johri P, Spitze K et al (2017) Estimating seven coefficients of pairwise relatedness using population-genomic data. Genetics. https://doi.org/10.1534/genetics.116.190660
Artyukova EV, Kozyrenko MM, Kholina AB et al (2011) High chloroplast haplotype diversity in the endemic legume Oxytropis chankaensis may result from independent polyploidization events. Genetica. https://doi.org/10.1007/s10709-010-9539-8
Bolger AM, Marc L, Bjoern U (2014) Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. https://doi.org/10.1093/bioinformatics/btu170
Bordbar F, Mohammadabadi M, Jensen J et al (2022) Identification of candidate genes regulating carcass depth and hind leg circumference in simmental beef cattle using Illumina Bovine Beadchip and next-generation sequencing analyses. Animals. https://doi.org/10.3390/ani12091103
Bradbury PJ, Zhang Z, Kroon DE et al (2007) TASSEL: Software for association mapping of complex traits in diverse samples. Bioinformatics. https://doi.org/10.1093/bioinformatics/btm308
Brunton AJ, Lamont RW, Conroy GC et al (2022) Phylogenetic reconstruction of the rainforest lineage Fontainea heckel (Euphorbiaceae) based on chloroplast DNA sequences and reduced-representation SNP Markers. Diversity. https://doi.org/10.3390/d14090725
Burns MJ, Edwards KJ, Newbury HJ et al (2001) Development of simple sequence repeat (SSR) markers for the assessment of gene flow and genetic diversity in pigeonpea (Cajanus cajan). Mol Ecol Notes. https://doi.org/10.1046/j.1471-8278.2001.00109.x
Carlson CS, Eberle MA, Rieder MJ et al (2004) Selecting a maximally informative set of single-nucleotide polymorphisms for association analyses using linkage disequilibrium. Am J Hum Genet. https://doi.org/10.1086/381000
Cay SB, Cinar YU, Kuralay SC et al (2022) Genome skimming approach reveals the gene arrangements in the chloroplast genomes of the highly endangered Crocus L. species: Crocus istanbulensis (B. Mathew) Rukšāns. PLoSOne. https://doi.org/10.1371/journal.pone.0269747
Chacón SMI, Pickersgill B, Debouck DG (2005) Domestication patterns in common beans (Phaseolus vulgaris L.) and the origin of the Mesoamerican and Andean cultivated races. Theoret Appl Genet. https://doi.org/10.1007/s00122-004-1842-2
Cingolani P, Platts A, Wang LL et al (2012) A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strain w1118; iso-2; iso-3. Fly. https://doi.org/10.4161/fly.19695
Cleeg MT, Zurawski G (1992) Chloroplast DNA and the study of plant phylogeny. New York, In: Soltis PS, Soltis DE, Doyle JJ (Eds.) Present status and future prospects in Molecular Systematics of Plants, Chapman & Hall. https://doi.org/10.1007/978-1-4615-3276-7_1
Cook DE, Andersen EC (2017) VCF-kit: assorted utilities for the variant call format. Bioinformatics. https://doi.org/10.1093/bioinformatics/btx011
Datta S, Singh P, Mahfooz S et al (2016) Novel genic microsatellite markers from Cajanus scarabaeoides and their comparative efficiency in revealing genetic diversity in pigeonpea. J Genet. https://doi.org/10.1007/s12041-013-0230-6
deJong MJ, deJong JF, Hoelzel AR et al (2021) SambaR: An R package for fast, easy and reproducible population genetic analyses of biallelic SNP data sets. Mol Ecol Resour. https://doi.org/10.1111/1755-0998.13339
Desiderio F, Bitocchi E, Bellucci E et al (2013) Chloroplast microsatellite diversity in Phaseolus vulgaris. Front Plant Sci. https://doi.org/10.3389/fpls.2012.00312
Doebley JF, Brandon S, Bruce SD (2006) The molecular genetics of crop domestication. Cell. https://doi.org/10.1016/j.cell.2006.12.006
Earl DA (2012) Structure harvester: a website and program for visualizing structure output and implementing the Evanno method. Conserv Genet Resour. https://doi.org/10.1007/s12686-011-9548-7
Erixon P, Oxelman B (2008) Whole-gene positive selection, elevated synonymous substitution rates, duplication, and indel evolution of the chloroplast clpP1 gene. PLoS One. https://doi.org/10.1371/journal.pone.0001386
Evanno GS, Regnaut S, Goudet J (2005) Detecting the number of clusters of individuals using the software structure: a simulation study. Mol Ecol. https://doi.org/10.1111/j.1365-294X.2005.02553.x
Falush D, Stephens M, Pritchard JK (2003) Inference of population structure using multilocus genotype data: linked loci and correlated allele frequencies. Genetics. https://doi.org/10.1093/genetics/164.4.1567
Garrison E, Gabor M (2012) Haplotype-based variant detection from short-read sequencing. arXiv. https://doi.org/10.48550/arXiv.1207.3907
Gaut B, Yang L, Takuno S et al (2011) The patterns and causes of variation in plant nucleotide substitution rates. Annu Rev Ecol Evol Syst. https://doi.org/10.1146/annurev-ecolsys-102710-145119
Goudet J (2005) Hierfstat, a package for R to compute and test hierarchical F statistics. Mol Ecol Notes. https://doi.org/10.1111/j.1471-8286.2004.00828.x
Honorio Coronado EN, Blanc-Jolivet C, Mader M et al (2020) SNP markers as a successful molecular tool for assessing species identity and geographic origin of trees in the economically important South American legume genus Dipteryx. J Hered. https://doi.org/10.1093/jhered/esaa011
Hughes AR, Inouye BD, Johnson MT et al (2008) Ecological consequences of genetic diversity. Ecol Lett. https://doi.org/10.1111/j.1461-0248.2008.01179.x
Jombart T (2008) adegenet: a R package for the multivariate analysis of genetic markers. Bioinformatics. https://doi.org/10.1093/bioinformatics/btn129
Jombart T, Ahmed I (2011) Adegenet 1.3–1: new tools for the analysis of genome-wide SNP data. Bioinformatics. https://doi.org/10.1093/bioinformatics/btr521
Kassa MT, Penmetsa RV, Carrasquilla-Garcia N et al (2012) Genetic patterns of domestication in pigeonpea (Cajanus cajan (L.) Millsp.) and wild Cajanus relatives. PloS One. https://doi.org/10.1371/journal.pone.0039563
Keenan K, McGinnity P, Cross TF et al (2013) diveRsity: An R package for the estimation and exploration of population genetics parameters and their associated errors. Methods Ecol Evol. https://doi.org/10.1111/2041-210X.12067
Keller J, Rousseau-Gueutin M, Martin GE et al (2017) The evolutionary fate of the chloroplast and nuclear rps16 genes as revealed through the sequencing and comparative analyses of four novel legume chloroplast genomes from Lupinus. DNA Res. https://doi.org/10.1093/dnares/dsx006
Kilian B, ÖzkanH WA et al (2007) Molecular diversity at 18 loci in 321 wild and 92 domesticate lines reveal no reduction of nucleotide diversity during Triticum monococcum (einkorn) domestication: implications for the origin of agriculture. Mol Biol Evol. https://doi.org/10.1093/molbev/msm192
Kimaro D, Melis R, Sibiya J (2020) Analysis of genetic diversity and population structure of Pigeonpea [Cajanus cajan (L.) Millsp] accessions using SSR markers. Plants. https://doi.org/10.3390/plants9121643
Kimura M (1983) Rare variant alleles in the light of the neutral theory. Mol Biol Evol. https://doi.org/10.1093/oxfordjournals.molbev.a040305
Knaus BJ, Grünwald NJ (2017) vcfR: a package to manipulate and visualize variant call format data in R. Mol Ecol Resour. https://doi.org/10.1111/1755-0998.12549
Konopiński MK (2020) Shannon diversity index: a call to replace the original Shannon’s formula with unbiased estimator in the population genetics studies. PeerJ. https://doi.org/10.7717/peerj.9391
Kumar S, Stecher G, Li M et al (2018) MEGA X: molecular evolutionary genetics analysis across computing platforms. Mol Biol Evol. https://doi.org/10.1093/molbev/msy096
Kumar K, Anjoy P, Sahu S et al (2022) Single trait versus principal component based association analysis for flowering related traits in pigeonpea. Sci Rep. https://doi.org/10.1038/s41598-022-14568-1
Kumar K, Bhattacharjee S, Vaikuntapu PR et al (2020) Climate change mitigation and adaptation through biotechnological interventions In Climate Change and Indian Agriculture: Challenges and Adaptation Strategies (Eds Ch. Srinivasarao et al.), ICAR-National Academy of Agricultural Research Management, India, pp 1–22
Li H, Bob H, Alec W et al (2009) The sequence alignment/map format and SAMtools. Bioinformatics. https://doi.org/10.1093/bioinformatics/btp352
Li B, Lin F, Huang P et al (2020a) Development of nuclear SSR and chloroplast genome markers in diverse Liriodendron chinense germplasm based on low-coverage whole genome sequencing. Biol Res. https://doi.org/10.1186/s40659-020-00289-0
Li C, Zheng Y, Huang P (2020b) Molecular markers from the chloroplast genome of rose provide a complementary tool for variety discrimination and profiling. Sci Rep. https://doi.org/10.1038/s41598-020-68092-1
Li QJ, Su N, Zhang L et al (2020c) Chloroplast genomes elucidate diversity, phylogeny, and taxonomy of Pulsatilla (Ranunculaceae). Sci Rep. https://doi.org/10.1038/s41598-020-76699-7
Li S, Liu SL, Pei SY et al (2020d) Genetic diversity and population structure of Camellia huana (Theaceae), a limestone species with narrow geographic range, based on chloroplast DNA sequence and microsatellite markers. Plant Diversity. https://doi.org/10.1016/j.pld.2020.06.003
Li H (2013) Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM. arXiv. https://doi.org/10.48550/arXiv.1303.3997
Lynch M, Ritland K (1999) Estimation of pairwise relatedness with molecular markers. Genetics. https://doi.org/10.1093/genetics/152.4.1753
Mader M, Pakull B, Blanc-Jolivet C et al (2018) Complete chloroplast genome sequences of four Meliaceae species and comparative analyses. Int J Mol Sci. https://doi.org/10.3390/ijms19030701
Van der Maesen LJG (1991) Pigeonpea: origin, history, evolution and taxonomy. In The pigeonpea (Eds Nene et al.), CAB, pp. 15–46
Maréchal A, Brisson N (2010) Recombination and the maintenance of plant organelle genome stability. New Phytol. https://doi.org/10.1111/j.1469-8137.2010.03195.x
Mohammadabadi MR, Esfandyarpoor E, Mousapour A (2017) Using inter simple sequence repeat multi-loci markers for studying genetic diversity in Kermani sheep. J Res Develop 5(2):e154
Muse SV, Gaut BS (1994) A likelihood approach for comparing synonymous and nonsynonymous nucleotide substitution rates, with application to the chloroplast genome. Mol Biol Evol. https://doi.org/10.1093/oxfordjournals.molbev.a040152
Nei M, Li WH (1979) Mathematical model for studying genetic variation in terms of restriction endonucleases. Proc Natl Acad Sci. https://doi.org/10.1073/pnas.76.10.5269
Notter DR (1999) The importance of genetic diversity in livestock populations of the future. J Anim Sci. https://doi.org/10.4236/ajps.2013.411264
Pandey P, Kumar R, Pandey VR et al (2013) Genetic divergence studies in pigeonpea [Cajanus cajan (L.) Millsp]. Am J Plant Sci. https://doi.org/10.4236/ajps.2013.411264
Peakall R, Smouse PE (2006) GENALEX 6: genetic analysis in Excel. Population genetic software for teaching and research. Mol Ecol Notes. https://doi.org/10.1111/j.1471-8286.2005.01155.x
Peakall R, Smouse PE (2012) GenAlEx 6.5: genetic analysis in Excel. Population genetic software for teaching and research-an update. Bioinformatics. https://doi.org/10.1093/bioinformatics/bts460
Pervaiz T, Zhang C, Faheem M (2017) Chloroplast based genetic diversity among Chinese grapes genotypes. Mitochondrial DNA Part A. https://doi.org/10.3109/24701394.2016.1155119
Petit RJ, Duminil J, Fineschi S (2005) Comparative organization of chloroplast, mitochondrial and nuclear diversity in plant populations. Mol Ecol. https://doi.org/10.1111/j.1365-294X.2004.02410.x
Pritchard JK, Stephens P, Donnelly P (2000) Inference of population structure using multi-locus genotype data. Genetics. https://doi.org/10.1093/genetics/155.2.945
Purseglove JW (1976) The origins and migrations of crops in tropical Africa. In: Harlan JR, De Wet JMJ, Stemler ABL (eds) Origins of African plant domestication. Mouton Publishers, The Hague, pp 291–310
Ray Choudhury P, Singh IP, George B et al (2008) Assessment of genetic diversity of pigeonpea cultivars using RAPD analysis. Biol Plant. https://doi.org/10.1007/s10535-008-0126-0
Rekha R, Prasanthi L, Sekhar MR et al (2011) Genetic diversity in pigeonpea [Cajanus cajan (L.) Millsp]. Legume Res 34(2):139–142
Rockenbach K, Havird JC, Monroe JG et al (2016) Positive selection in rapidly evolving plastid–Nuclear enzyme complexes. Genetics. https://doi.org/10.1534/genetics.116.188268
Sharma S, Kumar K, Tribhuvan K et al (2022) High throughput genotyping platforms in genotyping by sequencing for crop improvement (Eds Sonah et al.). John Wiley & Sons. https://doi.org/10.1002/9781119745686.ch2
Shaw J, Lickey EB, Beck JT et al (2005) The tortoise and the hare II: relative utility of 21 noncoding chloroplast DNA sequences for phylogenetic analysis. Am J Bot. https://doi.org/10.3732/ajb.92.1.142
Shen P, Wang F, Underhill PA et al (2000) Population genetic implications from sequence variation in four Y chromosome genes. Proc Natl Acad Sci. https://doi.org/10.1073/pnas.97.13.7354
Skuza L, Szućko I, Filip E et al (2019) Genetic diversity and relationship between cultivated, weedy and wild rye species as revealed by chloroplast and mitochondrial DNA non-coding regions analysis. PLoS One. https://doi.org/10.1371/journal.pone.0213023
Skuza L, Filip E, Szućko I (2013) Use of organelle markers to study genetic diversity in soybean. In a comprehensive survey of international soybean research-genetics, physiology, agronomy and nitrogen relationships (Eds James Board). IntechOpen. https://doi.org/10.5772/52028
Sloan DB, Triant DA, Forrester NJ et al (2014) A recurring syndrome of accelerated plastid genome evolution in the angiosperm tribe Sileneae (Caryophyllaceae). Mol Phylogenet Evol. https://doi.org/10.1016/j.ympev.2013.12.004
Songok S, Ferguson M, Muigai AW et al (2010) Genetic diversity in pigeonpea [Cajanus cajan (L.) Millsp] landraces as revealed by simple sequence repeat markers. Afr J Biotechnol 9(22):3231–3241
Tajima F (1989) Statistical method for testing the neutral mutation hypothesis by DNA polymorphism. Genetics. https://doi.org/10.1093/genetics/123.3.585
Turakulov R, Easteal S (2003) Number of SNPs loci needed to detect population structure. Hum Hered. https://doi.org/10.1159/000071808
Varshney RK, Chen W, Li Y et al (2012) Draft genome sequence of pigeonpea (Cajanuscajan), an orphan legume crop of resource-poor farmers. Nat Biotechnol. https://doi.org/10.1038/nbt.2022
Varshney RK, Saxena RK, Upadhyaya HD et al (2017) Whole-genome resequencing of 292 pigeonpea accessions identifies genomic regions associated with domestication and agronomic traits. Nat Genet 49(7):1082
Visser C, Lashmar SF, Van Marle-Köster E et al (2016) Genetic diversity and population structure in South African, French and Argentinian Angora goats from genome-wide SNP data. PLoS One. https://doi.org/10.1371/journal.pone.0154353
von Thaden A, Nowak C, Tiesmeyer A et al (2020) Applying genomic data in wildlife monitoring: Development guidelines for genotyping degraded samples with reduced single nucleotide polymorphism panels. Mol Ecol Resour. https://doi.org/10.1111/1755-0998.13136
Watterson GA (1975) On the number of segregating sites in genetical models without recombination. Theor Popul Biol. https://doi.org/10.1016/0040-5809(75)90020-9
Xiao-Ming Z, Junrui W, Li F et al (2017) Inferring the evolutionary mechanism of the chloroplast genome size by comparing whole-chloroplast genome sequences in seed plants. Sci Rep. https://doi.org/10.1038/s41598-017-01518-5
Yan M, Xiong Y, Liu R et al (2018) The application and limitation of universal chloroplast markers in discriminating east Asian evergreen oaks. Front Plant Sci. https://doi.org/10.3389/fpls.2018.00569
Yang S, Pang W, Ash G et al (2006) Low level of genetic diversity in cultivated Pigeonpea compared to its wild relatives is revealed by diversity arrays technology. Theoret Appl Genet. https://doi.org/10.1007/s00122-006-0317-z
Zavinon F, Adoukonou-Sagbadja H, Keilwagen J et al (2020) Genetic diversity and population structure in Beninese pigeonpea [Cajanus cajan (L.) Huth] landraces collection revealed by SSR and genome wide SNP markers. Genet Resour Crop Evolut. https://doi.org/10.1007/s10722-019-00864-9
Zehdi-Azouzi S, Cherif E, Moussouni S et al (2015) Genetic structure of the date palm (Phoenix dactylifera) in the Old World reveals a strong differentiation between eastern and western populations. Ann Bot. https://doi.org/10.1093/aob/mcv068
Zeng K, Fu YX, Shi S et al (2006) Statistical tests for detecting positive selection by utilizing high frequency variants. Genetics. https://doi.org/10.1534/genetics.106.061432
Zhu A, GuoW JK et al (2014) Unprecedented heterogeneity in the synonymous substitution rate within a plant genome. Mol Biol Evol. https://doi.org/10.1093/molbev/msu079
Acknowledgements
We acknowledge the support provided by the Director, ICAR-NIPB and PG School ICAR-IARI, New Delhi, India. The research work carried out in this manuscript is a major part of KK’s Ph.D thesis and was funded majorly by ICAR-IARI (NAHEP-CAAST). KK also acknowledged the CSIR-UGC for the fellowship received during his Ph.D. The authors are grateful and acknowledge the support provided by the ICAR-IARI (NAHEP-CAAST) program.
Funding
The present manuscript does not involve any type/source of funding.
Author information
Authors and Affiliations
Contributions
KK, SS and KG have designed the whole study. KK, PG, KNS and HS together performed the data analysis. KK and MSN conducted the CAPS marker development and validation experiments. KK, AM, SS, PKJ, KD, and KG interpreted the results obtained. KK, SS, AM, KD, KG and PKJ wrote the manuscript. All authors have read and approved the final manuscript.
Corresponding author
Ethics declarations
Competing interests
Authors declare that no significant competing financial, professional or personal interests that might have influenced the performance or presentation of the work described in this manuscript exists.
Ethical approval
This article does not include any study with humans or animals.
Consent to participate
Not applicable.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.
About this article
Cite this article
Kumar, K., Gupta, P., Singh, K.N. et al. Whole chloroplast genome-specific non-synonymous SNPs reveal the presence of substantial diversity in the pigeonpea mini-core collection. 3 Biotech 13, 365 (2023). https://doi.org/10.1007/s13205-023-03785-8
Received:
Accepted:
Published:
DOI: https://doi.org/10.1007/s13205-023-03785-8
Keywords
Profiles
- Kishor Gaikwad View author profile