Abstract
Key message
We report here that the mutation causing fractured starch granules in the barley line “Franubet” results from a point mutation in the barley gene corresponding to the rice FLO6 gene.
Abstract
The “fra” mutation in barley, which was originally isolated and characterized over 30 years ago, results in fractured starch granules and an opaque phenotype. This mutation has been used in breeding programs, since it appears to be useful in the production of pearled barley for human consumption. However, selection for this phenotype is difficult, since wild-type and heterozygous kernels cannot be distinguished phenotypically, and until now, the gene involved in this mutation has not been determined. Here, we used a map-based cloning approach using nanopore sequencing to obtain long reads from a BAC clone carrying markers on either side of the fra locus. By fine mapping followed by aligning RNA-seq reads to four genes within the mapped region, we were able to determine that the fra mutation is caused by the introduction of a stop codon in the barley homologue of the rice FLOURY ENDOSPERM 6 (FLO6) gene. This gene has a CBM48 domain that binds to starch, and may act through interactions with isoamylase1 (ISA1), assisting in the binding of ISA1 to starch granules. Perfect markers able to distinguish all genotypes were designed and tested in several large populations; in all cases, the markers were able to distinguish wild-type, heterozygous, and mutant genotypes.
Similar content being viewed by others
References
Altschule SF, Madden TL, Schaeffer AA (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402
Bahaji A, Li J, Sánchez-López ÁM, Baroja-Fernández E, Muñoz FJ, Ovecka M, Almagro G, Montero M, Ezquer I, Etxeberria E, Pozueta-Romero J (2014) Starch biosynthesis, its regulation and biotechnological approaches to improve crop yields. Biotechnol Adv 32:87–106
Baik BK, Ullrich SE (2008) Barley for food: characteristics, improvement, and renewed interest. J Cereal Sci 48:233–242
Ball S, Guan HP, James M, Myers A, Keeling P, Mouille G, Buléon A, Colonna P, Preiss J (1996) From glycogen to amylopectin: a model for the biogenesis of the plant starch granule. Cell 86:349–352
Beier S, Himmelbach A, Colmsee C, Zhang XQ, Barrero RA, Zhang Q, Li L, Bayer M, Bolser D, Taudien S, Groth M (2017) Construction of a map-based reference genome sequence for barley, Hordeum vulgare L. Sci Data 4:170044
Biggerstaff D (1988) Utilizing lethal translocation homozygotes of barley (Hordeum vulgare L.). Doctoral dissertation, Montana State University-Bozeman, College of Agriculture
Burton RA, Jenner H, Carrangis L, Fahy B, Fincher GB, Hylton C, Laurie DA, Parker M, Waite D, Van Wegen S, Verhoeven T (2002) Starch granule initiation and growth are altered in barley mutants that lack isoamylase activity. Plant J 31:97–112
Cantarel BL, Coutinho PM, Rancurel C, Bernard T, Lombard V, Henrissat B (2008) The Carbohydrate-Active EnZymes database (CAZy): an expert resource for glycogenomics. Nucleic Acids Res 37:D233–D238
Chung TY (1982) Isolation, description, inheritance, associated traits and possible uses of three barley (Hordeum vulgare L.) starch mutants. Doctoral dissertation, Montana State University-Bozeman, College of Agriculture
Chung TY (2001) lnheritence, linkage and possible use of fractured starch mutant in barley (Hordeum vulgare L.). J Plant Biotechnol 3:151–157
Colmsee C, Beier S, Himmelbach A, Schmutzer T, Stein N, Scholz U, Mascher M (2015) BARLEX—the barley draft genome explorer. Mol Plant 8:964–966
Edney MJ, Rossnagel BG, Endo Y, Ozawa S, Broph M (2002) Pearling quality of Canadian barley varieties and their potential use as rice extenders. J Cereal Sci 36:295–305
Evers AD, O’Brien L, Blakeney AB (1999) Cereal structure and composition. Aust J Agr Res 50:629–650
Hallauer AR (2000) Specialty corns, 2nd edn. CRC Press, New York
Hardie DG, Hawley SA, Scott JW (2006) AMP-activated protein kinase–development of the energy sensor concept. J Physio 574:7–15
Hayden MJ, Tabone T, Mather DE (2009) Development and assessment of simple PCR markers for SNP genotyping in barley. Theor Appl Genet 119:939–951
Hearnden PR, Eckermann PJ, McMichael GL, Hayden MJ, Eglinton JK, Chalmers KJ (2007) A genetic map of 1,000 SSR and DArT markers in a wide barley cross. Thero Appl Genet 115:383
International Barley Genome Sequencing Consortium (2012) A physical, genetic and functional sequence assembly of the barley genome. Nature 491:711–716
Kaur KD, Jha A, Sabikhi L, Singh AK (2014) Significance of coarse cereals in health and nutrition: a review. J Food Sci Technol 51:1429–1441
Kawagoe Y, Kubo A, Satoh H, Takaiwa F, Nakamura Y (2005) Roles of isoamylase and ADP-glucose pyrophosphorylase in starch granule synthesis in rice endosperm. Plant J 42:164–174
Kosambi DD (1944) Efficient mapping of a female sterile gene in wheat (Triticum aestivum L.). Ann Eugenics 12:172–175
Kubo A, Fujita N, Harada K, Matsuda T, Satoh H, Nakamura Y (1999) The starch-debranching enzymes isoamylase and pullulanase are both involved in amylopectin biosynthesis in rice endosperm. Plant Physiol 121:399–410
Liu F, Ahmed Z, Lee EA, Donner E, Liu Q, Ahmed R, Morell MK, Emes MJ, Tetlow IJ (2012) Allelic variants of the amylose extender mutation of maize demonstrate phenotypic variation in starch structure resulting from modified protein–protein interactions. J Exp Bot 63:1167–1183
Lloyd JR, Kossmann J (2015) Transitory and storage starch metabolism: two sides of the same coin? Curr Opin Biotechnol 32:143–148
Lorieux M (2012) MapDisto: fast and efficient computation of genetic linkage maps. Mol Breed 30:1231–1235
Mascher M et al (2017) A chromosome conformation capture ordered sequence of the barley genome. Nature 544(7651):427–433
McBride A, Ghilagaber S, Nikolaev A, Hardie DG (2009) The glycogen-binding domain on the AMPK β subunit allows the kinase to act as a glycogen sensor. Cell Metab 9:23–34
Modjtahedi N, Tokatlidis K, Dessen P, Kroemer G (2016) Mitochondrial proteins containing coiled-coil-helix-coiled-coil-helix (CHCH) domains in health and disease. Trends Biochem Sci 41:245–260
Muñoz-Amatriaín et al (2015) Sequencing of 15,622 gene-bearing BACs reveals new features of the barley genome. Plant J 84:216–227
Myers AM, Morell MK, James MG, Ball SG (2000) Recent progress toward understanding biosynthesis of the amylopectin crystal. Plant Physiol 122:989–998
Nakamura T, Vrinten P, Shimbata T, Saito M (2015) Starch modification: a model for wheat MAS breeding. In: Ogihara Y, Takumi S, Handa H (eds) Advances in wheat genetics: from genome to field. Springer, Tokyo, pp 265–273
Numa H, Itoh T (2014) MEGANTE: a web-based system for integrated plant genome annotation. Plant Cell Physiol 55(1):e2
Peng C, Wang Y, Liu F, Ren Y, Zhou K, Lv J, Zheng M, Zhao S, Zhang L, Wang C, Jiang L (2014) FLOURY ENDOSPERM6 encodes a CBM48 domain-containing protein involved in compound granule formation and starch synthesis in rice endosperm. Plant J 77:917–930
Polekhina G, Gupta A, Michell BJ, van Denderen B, Murthy S, Feil SC, Jennings IG, Campbell DJ, Witters LA, Parker MW, Kemp BE (2003) AMPK β subunit targets metabolic stress sensing to glycogen. Curr Biol 13:867–871
Sakai H, Lee SS, Tanaka T et al (2013) Rice Annotation Project Database (RAP-DB): an integrative and interactive database for rice genomics. Plant Cell Physiol 54:e6
Smith AM, Denyer K, Martin CR (1995) What controls the amount and structure of starch in storage organs? Plant Phys 107:673–677
Song Y, Jane J (2000) Characterization of barley starches of waxy, normal, and high amylose varieties. Carbohydr Polym 41:365–377
Stein N, Prasad M, Scholz U, Thiel T, Zhang H, Wolf M, Kota R, Varshney RK, Perovic D, Grosse I, Graner A (2007) A 1,000-loci transcript map of the barley genome: new anchoring points for integrative grass genomics. Thero Appl Genet 114:823–839
Suh DS, Verhoeven T, Denyer K, Jane JL (2004) Characterization of Nubet and Franubet barley starches. Carbohydr Polym 56:85–93
Tonooka T, Kawada N, Fujita M, Yoshikawa T, Kiribuch-Otobe C (2010) Usefulness of starch mutants in improvement of pearling quality of barley cultivated in an andosol upland field. Jpn J Crop Sci 79:308–315 (in Japanese with English abstract)
Toyosawa Y, Kawagoe Y, Matsushima R, Crofts N, Ogawa M, Fukuda M, Kumamaru T, Okazaki Y, Kusano M, Saito K, Toyooka K (2016) Deficiency of starch synthase IIIa and IVb alters starch granule morphology from polyhedral to spherical in rice endosperm. Plant Phyiol 170:1255–1270
Vandeputte GE, Delcour JA (2004) From sucrose to starch granule to starch physical behaviour: a focus on rice starch. Carbohydr Polym 58:245–266
Varshney RK, Marcel TC, Ramsay L, Russell J, Röder MS, Stein N, Waugh R, Langridge P, Niks RE, Graner A (2007) A high density barley microsatellite consensus map with 775 SSR loci. Thero Appl Genet 114:1091–1103
Verhoeven TM (2002) Determination of the morphology of starch granules in cereal endosperm. Doctoral dissertation, University of East Anglia
Wang YY, Tsay YF (2011) Arabidopsis nitrate transporter NRT1.9 is important in phloem nitrate transport. Plant Cell 23:1945–1957
Wang YJ, White PJ, Pollak LM, Jane J (1993) Characterization of starch structures of 17 maize endosperm mutant genotypes with Oh43 inbred line background. Cereal Chem 70:171–179
Acknowledgements
We would like to thank Drs. Masoko Seki, Tonooka Takuji and Takashi Yanagisawa for providing plant materials and suggestions for the work. We also thank Dr. Goro Ishikawa for his suggestions regarding preparation of the genetic maps.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors declare that they have no conflict of interest.
Additional information
Communicated by Ian D. Godwin.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Saito, M., Tanaka, T., Sato, K. et al. A single nucleotide polymorphism in the “Fra” gene results in fractured starch granules in barley. Theor Appl Genet 131, 353–364 (2018). https://doi.org/10.1007/s00122-017-3006-1
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00122-017-3006-1