Abstract
Citrus is a source of nutritional and medicinal advantages, cultivated worldwide with major groups of sweet oranges, mandarins, grapefruits, kumquats, lemons and limes. Pakistan produces all major citrus groups with mandarin (Citrus reticulata) being the prominent group that includes local commercial cultivars Feutral’s Early, Dancy, Honey, and Kinnow. The present study designed to understand the genetic architecture of this unique variety of Citrus reticulata ‘Kinnow.’ The whole-genome resequencing and variant calling was performed to map the genomic variability that might be responsible for its particular characteristics like taste, seedlessness, juice content, thickness of peel, and shelf-life. A total of 139,436,350 raw sequence reads were generated with 20.9 Gb data in Fastq format having 98% effectiveness and 0.2% base call error rate. Overall, 3,503,033 SNPs, 176,949 MNPs, 323,287 INS, and 333,083 DEL were identified using the GATK4 variant calling pipeline against Citrus clementina. Furthermore, g:Profiler was applied for annotating the newly found variants, harbor genes/transcripts and their involved pathways. A total of 73,864 transcripts harbors 4,336,352 variants, most of the observed variants were predicted in non-coding regions and 1009 transcripts were found well annotated by different databases. Out of total aforementioned transcripts, 588 involved in biological processes, 234 in molecular functions and 167 transcripts in cellular components. In a nutshell, 18,153 high impact variants and 216 genic variants found in the current study, which may be used after its functional validation for marker-assisted breeding programs of “Kinnow” to propagate its valued traits for the improvement of contemporary citrus varieties in the region.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
NGS-binary alignment maps (bam) files are submitted at NCBI SRA project ID: PRJNA821664. Furthermore supplementary tables and figures are also provided along with the manuscript. The same manuscript is also posted as preprint doi:https://doi.org/10.1101/2022.12.07.519411. Complete data will be available from the corresponding author on request.
References
Ahmed K (2020) Third of Pakistan’s 2020 mandarin exports earmarked for the Middle East. Arab News PK https://arab.news/y4hrw.
Altaf S, Khan MM, Jaskani MJ, Khan IA, Usman M, Sadia B, Awan FS, Ali A, Khan AI (2014) Morphogenetic characterization of seeded and seedless varieties of Kinnow Mandarin (‘Citrus reticulata’ Blanco). Aust J Crop Sci 8:1542–1549 https://www.researchgate.net/publication/268152281
Baqir HA, Zeboon NH, Al-behadili AJP (2019) The role and importance of amino acids within plants: a review. Plant Arch 19(2):1402–1410
Barbhuiya AR, Khan ML, Dayanandan S (2022) Molecular phylogeny of Citrus species in the Eastern Himalayan region of Northeast India based on chloroplast and nuclear DNA sequence data. In: Kumar A, Choudhury B, Dayanandan S, Khan ML (eds) Molecular genetics and genomics tools in biodiversity conservation. Springer, Singapore, pp 185–201. https://doi.org/10.1007/978-981-16-6005-4-9
Bi X, Liao L, Deng L, Jin Z, Huang Z, Sun G, Xiong B, Wang Z (2022) Combined transcriptome and metabolome analyses reveal candidate genes involved in Tangor (Citrus reticulata × Citrus sinensis) fruit development and quality formation. Int J Mol Sci 23:5457. https://doi.org/10.3390/ijms23105457
Bohry D, Ramos HCC, Santos PHD, Boechat MSB, Arêdes FAS, Pirovani AAV, Pereira MG (2021) Discovery of SNPs and InDels in papaya genotypes and its potential for marker assisted selection of fruit quality traits. Sci Rep 11:1–8. https://doi.org/10.1038/s41598-020-79401-z
Britten RJ, Rowen L, Williams J, Cameron RA (2003) Majority of divergence between closely related DNA samples is due to InDels. Proc Natl Acad Sci U S A 100:4661–4665. https://doi.org/10.1073/pnas.0330964100
Caballero JC, Alonso R, Ibañez V, Terol J, Talon M, Dopazo J (2015) Phylogenetic analysis of 34 chloroplast genomes elucidates the relationships between wild and domestic species within the genus Citrus. Mol Biol Evol 8:2015–2035. https://doi.org/10.1093/molbev/msv082
Chen C, Bowman KD, Choi YA et al (2008) EST-SSR genetic maps for Citrus sinensis and Poncirus trifoliata. Tree Genet Genomes 4:1–10. https://doi.org/10.1007/s11295-007-0083-3
Ching A, Caldwell KS, Jung M et al (2002) SNP frequency, haplotype structure and linkage disequilibrium in elite maize inbred lines. BMC Genet 3:1–14. https://doi.org/10.1186/1471-2156-3-19
Cingolani P, Platts A, Wang LL, Coon M, Nguyen T, Wang L, Land SJ, Lu X, Ruden DM (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 6:80–92. https://doi.org/10.4161/fly.19695
Cooper AJ, Dorai T, Pinto JT, Denton TT (2022) The metabolic importance of the overlooked asparaginase II pathway. Anal Biochem 644:114084. https://doi.org/10.1016/j.ab.2020.114084
Distefano G, Las CG, Deng X, Chai L (2020) Citrus reproductive biology from flowering to fruiting. In: Gentile A, La MS, Deng Z (eds) The citrus genome. Compendium of Plant Genomes. Springer, Cham, pp 167–176. https://doi.org/10.1007/978-3-030-15308-3-9
Duru S, Hayran S, Gül A (2022) The analysis of competitiveness of Mediterranean countries in the world citrus trade. Mediterr Agric Sci 35:21–26. https://doi.org/10.29136/mediterranean.1012466
Etxeberria E, Gonzalez P, Achor D, Albrigo G (2009) Anatomical distribution of abnormally high levels of starch in HLB-affected Valencia orange trees. Mol Plant Pathol 74:76–83. https://doi.org/10.1016/j.pmpp.2009.09.004
Fang Q, Wang L, Yu H et al (2018) Development of species-specific InDel markers in citrus. Plant Mol Biol Rep 36:653–662. https://doi.org/10.1007/s11105-018-1111-1
Fernandez CT (2022) Making a pangenome using the iterative mapping approach. In: Edwards D (ed) Plant bioinformatics. Methods in Molecular Biology. Humana, New York, pp 259–271. https://doi.org/10.1007/978-1-0716-2067-0-14
Froelicher Y, Mouhaya W, Bassene JB et al (2011) New universal mitochondrial PCR markers reveal new information on maternal citrus phylogeny. Tree Genet Genomes 7:49–61. https://doi.org/10.1007/s11295-010-0314-x
Galili G (2002) New insights into the regulation and functional significance of lysine metabolism in plants. Annu Rev Plant Biol 53:27
Gao Q, Yue G, Li W, Wang J, Xu J, Yin Y (2012) Recent progress using high-throughput sequencing technologies in plant molecular breeding. J Integr Plant Biol 54:215–227 http://www.jipb.net, http://www.wileyonlinelibrary.com/journal/jipb
García LA, Luro F, Navarro L et al (2012) Comparative use of InDel and SSR markers in deciphering the interspecific structure of cultivated citrus genetic diversity: a perspective for genetic association studies. Mol Genet Genomics 287:77–94. https://doi.org/10.1007/s00438-011-0658-4
Garcia LA, Curk F, Snoussi TH et al (2013) A nuclear phylogenetic analysis: SNPs, InDels and SSRs deliver new insights into the relationships in the ‘true citrus fruit trees’ group (Citrinae, Rutaceae) and the origin of cultivated species. Ann Bot 111:1–19. https://doi.org/10.1093/aob/mcs227
Ghada B, Amel O, Aymen M et al (2019) Phylogenetic patterns and molecular evolution among ‘True citrus fruit trees’ group (Rutaceae family and Aurantioideae subfamily). Sci Hortic 253:87–98. https://doi.org/10.1016/j.scienta.2019.04.011
Gill K, Kumar P, Kumar A et al (2022) Comprehensive mechanistic insights into the citrus genetics, breeding challenges, biotechnological implications, and omics-based interventions. Tree Genet Genomes 18:1–26. https://doi.org/10.1007/s11295-022-01544-z
Goswami M, Attri K, Goswami I (2022) Applications of molecular markers in fruit crops: a review. Int J Econ Plants 9:121–126
Goto S, Yoshioka T, Ohta S, Kita M, Hamada H, Shimizu T (2016) Segregation and heritability of male sterility in populations derived from progeny of Satsuma mandarin. PloS One 11:e0162408. https://doi.org/10.1371/journal.pone.0162408
Hayat F, Nawaz KM, Zafar SA, Balal R et al (2017) Surface coating and modified atmosphere packaging enhances storage life and quality of ‘Kaghzi lime’. J Agric Sci Technol 19:1151–1160 http://jast.modares.ac.ir/article-23-863-en.html
Huang X, Wu W, Su L, Lv H et al (2022) Development and application of InDel markers linked to fruit-shape and peel-colour genes in Wax Gourd. Genes 13:1567. https://doi.org/10.3390/genes13091567
Hughes HK, Rowland ME, Onore CE et al (2022) Dysregulated gene expression associated with inflammatory and translation pathways in activated monocytes from children with autism spectrum disorder. Transl Psychiatry 12:1–9. https://doi.org/10.1038/s41398-021-01766-0
Imai A, Nonaka K, Kuniga T et al (2018) Genome-wide association mapping of fruit-quality traits using genotyping-by-sequencing approach in citrus landraces, modern cultivars, and breeding lines in Japan. Tree Genet Genomes 14:24. https://doi.org/10.1007/s11295-018-1238-0
Jaskani MJ, Kwon SW, Kim DH (2005) Comparative study on vegetative, reproductive and qualitative traits of seven diploid and tetraploid watermelon lines. Euphytica 145:259–268. https://doi.org/10.1007/s10681-005-1644-x
Jiang D, Ye QL, Wang FS, Li CAO (2010) The mining of citrus EST-SNP and its application in cultivar discrimination. Agricl Sci China 9:179–190. https://doi.org/10.1016/S1671-2927(09)60082-1
Jiang P, Zhu T, Liu J, Tao X, Xue Z et al (2022) Mitochondrial DNA variant spectrum and the association with chronic tic disorders. Eur J Neurol 29:3187–3196
Kamal GM, Anwar F, Hussain AI, Sarri N, Ashraf MY (2011) Yield and chemical composition of citrus essential oils as affected by drying pretreatment of peels. Int Food Res J 18:1275–1282
Khan IA, Kender WJ (2007) Citrus breeding: introduction and objectives. Citrus genetics, breeding and biotechnology. CABI, United Kingdom
Khan MN, Muhammad A, Abid M, Tehseen A, Naveeda A, Hayat F, Raza S, Abdullah J, Rafi Q (2020) Characterization and comparative nutritional study of three strains of Kinnow mandarin (Citrus reticulata Blanco). Asian J Agric Biol 8:299–307
Külheim C, Hui YS, Maintz J, Foley WJ, Moran GF (2009) Comparative SNP diversity among four Eucalyptus species for genes from secondary metabolite biosynthetic pathways. BMC Genomics 10:1–11. https://doi.org/10.1186/1471-2164-10-452
Lea PJ, Sodek L, Parry MA, Shewry PR, Halford NG (2007) Asparagine in plants. Ann Appl Biol 150:1–26. https://doi.org/10.1111/j.1744-7348.2006.00104.x
Li H, Durbin R (2010) Fast and accurate long-read alignment with Burrows–Wheeler transform. Bioinformatics 26:589–595. https://doi.org/10.1093/bioinformatics/btp698
Liu E, Li J, Ou S, Dong B, Yang B, Zhou Y (2022) The complete mitochondrial genome of Semblis atrata (Trichoptera:Phryganeidae). s 7:956–958. https://doi.org/10.1080/23802359.2022.2080595
Liu TJ, Zhou JJ, Chen FY, Gan ZM, Li YP, Zhang JZ, Hu CG (2018) Identification of the genetic variation and gene exchange between Citrus trifoliata and Citrus clementina. Biomolecules 8:182. https://doi.org/10.3390/biom8040182
Liu F, Movahedi A, Yang W et al (2023) The complete plastid genome and characteristics analysis of Achillea millefolium. Funct Integr Genomics 23:192. https://doi.org/10.1007/s10142-023-01121-0
Marbouty M, Koszul R (2022) Metagenomes binning using proximity-ligation data. In: Bicciato S, Ferrari F (eds) Hi-C data analysis. Methods in Molecular Biology. Humana, New York, pp 163–181. https://doi.org/10.1007/978-1-0716-1390-0-8
Mattia MR, Du D, Yu Q, Kahn T, Roose M, Hiraoka Y, Wang Y, Munoz P, Gmitter FG Jr (2022) Genome-wide association study of healthful flavonoids among diverse mandarin accessions. Plants 11:317. https://doi.org/10.3390/plants11030317
Mills RE, Luttig CT, Larkins CE, Beauchamp A, Tsui C, Pittard WS, Devine SE (2006) An initial map of insertion and deletion (INDEL) variation in the human genome. Genome Res 16(9):1182–1190. https://doi.org/10.1101/gr.4565806
Montalt R, Vives MC, Navarro L, Ollitrault P, Aleza P (2021) Parthenocarpy and self-incompatibility in Mandarins. Agronomy 11:2023. https://doi.org/10.3390/agronomy11102023
Murray MG, Thompson WF (1980) Rapid isolation of high molecular weight plant DNA. Nucleic Acids Res 8:4321–4326. https://doi.org/10.1093/nar/8.19.4321
Naqvi SAH, Wang J, Malik MT, Umar UUD, Hasnain A, Sohail MA, Shakeel MT, Nauman M, Hassan MZ, Fatima MJA (2022) Citrus canker-distribution, taxonomy, epidemiology, disease cycle, pathogen biology, detection, and management: a critical review and future research agenda. Agronomy 12(5):1075. https://doi.org/10.3390/agronomy12051075
Naz S, Shahzadi K, Rashid S, Saleem F, Zafarullah A, Ahmad S (2014) Molecular characterization and phylogenetic relationship of different citrus varieties of Pakistan. J Anim Plant Sci 24:315–320
Noda T, Daiou K, Mihara T et al (2022) Efficient method for generating citrus hybrids with polyembryonic Satsuma mandarin as the female parent. Mol 42:1–15. https://doi.org/10.1007/s11032-022-01324-6
Novelli VM, Takita MA, Machado MA (2004) Identification and analysis of single nucleotide polymorphisms (SNPs) in citrus. Euphytica 138:227–237. https://doi.org/10.1023/B:EUPH.0000047086.47988.82
Nuñez LG, Balladares C, Pavez C, Urra C, Sanhueza D, Vendramin E et al (2019) High-density genetic map and QTL analysis of soluble solid content, maturity date, and mealiness in peach using genotyping by sequencing. Sci Hortic 257:108734. https://doi.org/10.1016/j.scienta.2019.108734
Oliveira M, Azevedo L (2022) Molecular markers: an overview of data published for fungi over the last ten years. J Fungi 8:803. https://doi.org/10.3390/jof8080803
Ollitrault P, Terol JF, Chen C, Federici CT, et al (2011). A reference linkage map of C. clementina based on SNPs, SSRs and InDels:477. https://agritrop.cirad.fr/559981
Ollitrault F, Terol J, Martin AA, Pina JA, Navarro L, Talon M, Ollitrault P (2012) Development of indel markers from Citrus clementina (Rutaceae) BAC-end sequences and interspecific transferability in citrus. Am J Bot 99:e268–e273. https://doi.org/10.3732/ajb.1100569
Ollitrault P, Garcia-Lor A, Terol J, Curk F, Ollitrault F, Talón M, Navarro L (2015) Comparative values of SSRs, SNPs and InDels for citrus genetic diversity analysis. Acta Hortic 1065:457–466. https://doi.org/10.17660/ActaHortic.2015.1065.56
Oueslati A, Salhi-Hannachi A, Luro F, Vignes H, Mournet P, Ollitrault P (2017) Genotyping by sequencing reveals the interspecific C. maxima/C. reticulata admixture along the genomes of modern citrus varieties of mandarins, tangors, tangelos, orangelos and grapefruits. PloS One 12:e0185618. https://doi.org/10.1371/journal.pone.0185618
Panaro NJ, Yuen PKI, Sakazume T, Fortina P, Kricka LJ, Wilding P (2000) Evaluation of DNA fragment sizing and quantification by the Agilent 2100 Bioanalyzer. Clin Chem 46:1851–1853. https://doi.org/10.1093/clinchem/46.11.1851
Park S, Yu HJ, Mun JH et al (2010) Genome-wide discovery of DNA polymorphism in Brassica rapa. Mol Genet Genomics 283:135–145. https://doi.org/10.1007/s00438-009-0504-0
Prasad H, Thakur M, Gupta AK, Prasad D (2015) Effect of foliar application of 2, 4-D, urea and zinc sulphate on fruit drop, yield and fruit quality of Kinnow mandarin. Int J Bio-Resour Stress Manag 6:619–622
Ramadugu C, Pfeil BE, Keremane ML, Lee RF, Maureira-Butler IJ, Roose ML (2013) A six nuclear gene phylogeny of Citrus (Rutaceae) taking into account hybridization and lineage sorting. PloS One 8:e68410. https://doi.org/10.1371/journal.pone.0068410
Raman H, Raman R, Wood R et al (2006) Repetitive indel markers within the ALMT1 gene conditioning aluminium tolerance in wheat (Triticum aestivum L.). Mol 18:171–183. https://doi.org/10.1007/s11032-006-9025-2
Rather SA, Wang T, Liu H et al (2023) Characterization of the complete chloroplast genome of Dalbergia congesta (Fabaceae), an endangered legume endemic to the Nilgiri Hills of Tamil Nadu, India. Funct Integr Genomics 23:126. https://doi.org/10.1007/s10142-023-01047-7
Raudvere U, Kolberg L, Kuzmin I, Arak T et al (2019) g:Profiler: a web server for functional enrichment analysis and conversions of gene lists (2019 update). Nucleic Acids Res 47:W191–W198. https://doi.org/10.1093/nar/gkz369
Rehman A, Deyuan Z, Hussain I, Iqbal MS, Yang Y, Jingdong L (2018) Prediction of major agricultural fruits production in pakistan by using an econometric analysis and machine learning technique. Int J Fruit Sci 18:445–461. https://doi.org/10.1080/15538362.2018.1485536
Ros R, Muñoz BJ, Krueger S (2014) Serine in plants: biosynthesis, metabolism, and functions. Trends Plant Sci 19:564–569. https://doi.org/10.1016/j.tplants.2014.06.003
Sabir I (2010) Pakistan, the largest kinnow grower. Daily times. https://defence.pk/pdf/threads/pakistan-currently-the-largest-kinnow-grower.84281/
Sambrook J, Fritsch E, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor, New York
Sambrook J, Green MR (2012) Molecular cloning: A laboratory manual. Cold Spring Harbor, New York
Terol J, Naranjo MA, Ollitrault P et al (2008) Development of genomic resources for Citrus clementina: characterization of three deep-coverage BAC libraries and analysis of 46,000 BAC end sequences. BMC Genomics 9:1–12. https://doi.org/10.1186/1471-2164-9-423
Tiwari JK, Yerasu SD, Rai N et al (2022) Progress in marker-assisted selection to genomics-assisted breeding in tomato. Crit Rev Plant Sci 41:321–350. https://doi.org/10.1080/07352689.2022.2130361
Torre FDL, Cañas RA, Pascual MB, Avila C, Cánovas FM (2014) Plastidic aspartate aminotransferases and the biosynthesis of essential amino acids in plants. J Exp Bot 65:5527–5534. https://doi.org/10.1093/jxb/eru240
Usman M, Fatima B, Gillani KA, Khan MS, Khan MM (2008) Exploitation of potential target tissues to develop polyploids in citrus. Pak J Bot 40:1755–1766
Vasemäg A, Gross R, Palm D et al (2010) Discovery and application of insertion-deletion (INDEL) polymorphisms for QTL mapping of early life-history traits in Atlantic salmon. BMC Genomics 11:1–11. https://doi.org/10.1186/1471-2164-11-156
Vauquelin LN, Robiquet PJ (1806) The discovery of a new plant principle in Asparagus sativus. Ann Chim 57:14
Wang N, Li C, Kuang L, Wu X et al (2022) Pan-mitogenomics reveals the genetic basis of cytonuclear conflicts in citrus hybridization, domestication, and diversification. Proc Natl Acad Sci 119:e2206076119. https://doi.org/10.1073/pnas.2206076119
Wu GA, Prochnik S, Jenkins J et al (2014) Sequencing of diverse mandarin, pummelo and orange genomes reveals complex history of admixture during citrus domestication. Nat Biotechnol 32:656–662. https://doi.org/10.1038/nbt.2906
Wu GA, Terol J, Ibanez V et al (2018) Genomics of the origin and evolution of citrus. Nature 554:311–316. https://doi.org/10.1038/nature25447
Wu GA, Sugimoto C, Kinjo H et al (2021) Diversification of mandarin citrus by hybrid speciation and apomixis. Nat Commun 12:4377. https://doi.org/10.1038/s41467-021-24653-0
Yamasaki M, Tenaillon MI, Vroh BI et al (2005) A large-scale screen for artificial selection in maize identifies candidate agronomic loci for domestication and crop improvement. Plant Cell 17:2859–2872. https://doi.org/10.1105/tpc.105.037242
Zhao C, Wang F, Lian Y, Xiao H, Zheng J (2020) Biosynthesis of citrus flavonoids and their health effects. Crit Rev Food Sci Nutr 60:566–583. https://doi.org/10.1080/10408398.2018.1544885
Acknowledgements
Authors are obliged to the Emeritus Prof. Dr. Mikeal Roose, University of California, Riverside, USA and Dr. Subhas Hajeri, Director-Citrus Pest Detection Program, Tulare, CA, USA for reviewing and providing the valued suggestions to improve the manuscript.
Author information
Authors and Affiliations
Contributions
Sadia Jabeen performed sample collection, wet-lab experimentation, WG variant calling pipeline, methodology/data analysis and initial draft write-up. Rashid Saif, WG variant calling pipeline design, data analysis, interpretation of results and initial draft write-up. Shagufta Naz conceptualization of the project, reviewing, proof reading and overall supervision. Rukhama Haq contributed as co-supervision of wet-lab experimentation, reviewing and editing the draft. Akbar Hayat performed identification of plants for sample collection.
Corresponding author
Ethics declarations
Ethics approval
Not applicable.
Competing interest
The authors declare no competing interests.
Additional information
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
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
Jabeen, S., Saif, R., Haq, R. et al. Whole-genome sequencing and variant discovery of Citrus reticulata “Kinnow” from Pakistan. Funct Integr Genomics 23, 227 (2023). https://doi.org/10.1007/s10142-023-01153-6
Received:
Revised:
Accepted:
Published:
DOI: https://doi.org/10.1007/s10142-023-01153-6