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Comparison of Methods to Distinguish Diploid and Tetraploid Potato in Applied Diploid Breeding

  • Maher Alsahlany
  • Daniel Zarka
  • Joseph Coombs
  • David S. DouchesEmail author
Article
  • 85 Downloads

Abstract

Diploid (2n = 2x = 24) and tetraploid (2n = 4x = 48) germplasm are commonly used in potato breeding programs. Potato breeders need to efficiently and inexpensively differentiate between diploid and tetraploid progeny in dihaploid induction crosses as well as 2x-2x crosses where 2n gametes may occur in the parents. In this study, we compared the chloroplast count, genome-wide SNP genotyping and flow cytometry methods to determine ploidy. Twenty-eight clones were used as reference samples (14 diploid lines (2n = 2x = 24), 14 tetraploid (2n = 4x = 48) varieties and advanced breeding lines) to compare the three ploidy determination methods. The chloroplast count method was used to determine the ploidy level in the 28 reference samples, and 102 potato breeding lines derived from diploid (2x-2x) crosses. The Infinium 12 K V2 Potato Single Nucleotide Polymorphism (SNP) Array was used to examine the 28 reference samples and the 102 breeding lines. The results obtained from both chloroplasts counts and SNP genotyping techniques determined that there was a total of 84 diploid lines and 18 tetraploid lines. Flow cytometry was also used to determine ploidy level in a subset of 42 lines (28 reference lines set and 14 breeding lines). All three methods of ploidy determination (chloroplast counting, SNP genotyping and flow cytometry) agreed for all samples evaluated. These results demonstrate the usefulness of chloroplast counting as an efficient and inexpensive method for breeders to differentiate ploidy between diploid and tetraploid potato in applied breeding programs.

Keywords

Potato ploidy level Chloroplast count Single nucleotide polymorphism (SNP) Flow cytometry Diploid potato (2n = 2x = 24) Tetraploid potato (2n = 4x = 48) 

Resumen

El germoplasma diploide (2n = 2x = 24) y el tetraploide (2n = 4X = 48) se usan comúnmente en los programas de mejoramiento de la papa. Los fitomejoradores necesitan diferenciar eficiente y económicamente entre progenies diploide y tetraploide en cruzas de inducción de dihaploides así como en cruzas 2x-2x, donde los gamentos 2n pudieran presentarse en los progenitores. En este estudio comparamos el conteo de cloroplastos, la amplitud genómica de SNP y los métodos de citometría de flujo para determinar la ploidía. Se usaron 28 clones como muestras de referencia (14 líneas diploides (2n = 2x = 24), 14 variedades tetraploides (2n = 4x = 48) y clones avanzados) para comparar los tres métodos de determinación de ploidía. El método del conteo de cloroplastos se usó para determinar el nivel de ploidía en las 28 muestras de referencia, y 102 líneas de papa de mejoramiento derivadas de cruzas diploides (2x-2x). Se usó el arreglo de polimorfismo de nucleótido simple (SNP) de papa Infinium 12 K V2 para examinar las 28 muestras de referencia y los 102 clones avanzados. Los resultados que se obtuvieron de ambas técnicas, el conteo de cloroplastos y la determinación de genotipos por SNP, determinaron que hubo un total de 84 líneas diploides y 18 tetraploides. También se usó la citometría de flujo para determinar el nivel de ploidía en un subgrupo de 42 líneas (un juego de 28 líneas de referencia y 14 de mejoramiento). Los tres métodos para la determinación de ploidías (el conteo de cloroplastos, el genotipado por SNP, y la citometría de flujo) coincidieron para todas las muestras evaluadas. Estos resultados demuestran la utilidad del conteo de cloroplastos como un método eficiente y económico para los mejoradores para diferenciar ploidías entre papa diploide y tetraploide para aplicar en programas de mejoramiento.

Notes

Acknowledgments

This research was funded by USDA NIFA 2014-67013-22434 and the Higher Committee for Education Development in Iraq (HCED). The authors thank all Michigan State University Potato Breeding and Genetics Program members and especially Dr. Norma C. Manrique-Carpintero and Nick Garrity, who helped with chloroplast counting. The authors also thank the Kathiravetpillai Arumuganathan lab for processing leaf samples with flow cytometry to measure DNA content. The authors thank Dr. Linda Hanson (USDA/ARS Sugar Beet Pathology Lab) for the use of their microscope for chloroplast counts and Dr. Mitch McGrath (USDA/ARS) for reviewing the manuscript.

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Copyright information

© The Potato Association of America 2019

Authors and Affiliations

  1. 1.Department of Plant, Soil and Microbial SciencesMichigan State UniversityEast LansingUSA
  2. 2.Horticulture Department College of AgricultureAl-Qasim Green UniversityBabylonIraq

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