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A comparative study of ancient DNA isolated from charred pea (Pisum sativum L.) seeds from an Early Iron Age settlement in southeast Serbia: inference for pea domestication

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Abstract

The development of agriculture was a key turning point in human history, a central part of which was the evolution of new plant forms, domesticated crops. Grain legumes were domesticated in parallel with cereals and formed important dietary components of early civilizations. First domesticated in the Near East, pea has been cultivated in Europe since the Stone and Bronze Ages. In this study, we present a molecular analysis of ancient DNA (aDNA) extracted from carbonized pea seeds recovered from deposits at Hissar, in southeast Serbia, that date to the eleventh century B.C. Four selected chloroplast DNA loci (trnSG, trnK, matK and rbcL) amplified in six fragments of 128–340 bp with a total length of 1,329 bp were successfully recovered in order to distinguish between cultivated and wild gathered pea. Based on identified mutations, the results showed that genuine aDNA was analyzed. Moreover, DNA analysis resulted in placing the ancient sample at an intermediate position between extant cultivated [Pisum sativum L. and wild P. sativum subsp. elatius (Steven ex M. Bieb.) Asch. et Graebn.]. Consequently, based on a combination of morphological and molecular data, we concluded that the material represents an early domesticated pea. We speculate that Iron Age pea would be of colored flower and pigmented testa, similar to today’s fodder pea (P. sativum subsp. sativum var. arvense (L.) Poir.), possibly of winter type. This is the first report of successful aDNA extraction and analysis from any legume species thus far. The implications for pea domestication are discussed here.

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Acknowledgments

This work was supported by the projects 173005, 173030, 31016 and 31024 of the Ministry of Education and Science of the Republic of Serbia, and SEELEGUMES-168 within the EU programme SEE-ERA.NET. P.S. acknowledges funding from Grant Agency of Palacký University in Olomouc, IGA PrF-2013-003. The authors cordially thank Noel Ellis, Gérard Duc for useful suggestions, Milorad Stojić for providing archeobotanical material and Clarice Coyne for manuscript style improvement.

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Authors and Affiliations

  1. Department of Botany, Faculty of Science, Palacký University in Olomouc, Šlechtitelů 11, 783 71, Olomouc, Czech Republic

    Petr Smýkal

  2. Plant Molecular Biology Lab, Institute of Molecular Genetics and Genetic Engineering, University of Belgrade, P.O. Box 23, 11010, Belgrade, Serbia

    Živko Jovanović & Nemanja Stanisavljević

  3. Department of Biology and Ecology, Faculty of Sciences and Mathematics, University of Niš, Višegradska 33, 18000, Niš, Serbia

    Bojan Zlatković

  4. Department of Field and Vegetable Crops, Faculty of Agriculture, University of Novi Sad, Trg Dositeja Obradovića 8, 21000, Novi Sad, Serbia

    Branko Ćupina

  5. Institute of Field and Vegetable Crops, Maksima Gorkog 30, 21000, Novi Sad, Serbia

    Vuk Đorđević & Aleksandar Mikić

  6. Museum of Vojvodina, Dunavska 35, 21000, Novi Sad, Serbia

    Aleksandar Medović

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10722_2014_128_MOESM1_ESM.rtf

Supplementary material 1 Complete alignments of respective cpDNA fragments with indicated variable polymorphic sites. A) trnSG intergenic spacer alignment with underlined primer sequences, dash indicates gap of 289bp, B) trnK and matK sequences alignment with indicated gap of 420bp, C) rbcL sequences alignment (RTF 97 kb)

10722_2014_128_MOESM2_ESM.docx

Supplementary material 2 Amplification success in each of two extractions indicated for respective cpDNA regions, ordered by increasing length from 128 to 599 bp fragments (symbolized by black triangle above). Numbers indicate number of the successful out of all performed PCR amplifications. Numbers by markers indicate the respective fragment length. Light grey indicate amplification success less than 50% and darker grey failure (0%) amplification (DOCX 19 kb)

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Smýkal, P., Jovanović, Ž., Stanisavljević, N. et al. A comparative study of ancient DNA isolated from charred pea (Pisum sativum L.) seeds from an Early Iron Age settlement in southeast Serbia: inference for pea domestication. Genet Resour Crop Evol 61, 1533–1544 (2014). https://doi.org/10.1007/s10722-014-0128-z

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