Abstract
Perennial ryegrass is an important agricultural crop, however, it is susceptible to winterkill. Freezing injury is caused primarily by ice formation. The LpIRI1 protein has the potential to inhibit ice recrystallization, thus minimize the damage. An association study was conducted using single nucleotide polymorphisms obtained through allele sequencing of the LpIRI1 gene and phenotypic data were collected using two phenotyping platforms in a perennial ryegrass association mapping population of 76 diverse genotypes. Winter survival (FWS) was evaluated under field conditions, while tiller survival (PTS) and electrolyte leakage (EL) at −8 and −12 °C were determined under controlled-environment conditions. Proline content (PC) in cold-acclimated plants was measured prior to the freezing test. Significant variation in FWS, PTS, EL and PC was observed among genotypes in our panel. EL and PTS revealed significant negative correlations at −8 °C (rs = −0.40) and −12 °C (rs = −0.49). PC, however, did not show significant correlations with any of the measured traits, while FWS was correlated (rs = −0.48) with EL at −12 °C. The LpIRI1 gene was found to be highly polymorphic with an average SNP frequency of 1 SNP per 16 bp. Association analysis revealed two non-synonymous SNPs being associated with increased EL, both located in the LpIRI1 leucine-rich repeat. The results indicate that allelic variation in the LpIRI1 gene plays an important role in the cell membrane integrity of perennial ryegrass during freezing, and can be exploited for developing more freezing tolerant cultivars.
Similar content being viewed by others
References
Ábrahám E, Hourton-Cabassa C, Erdei L, Szabados L (2010) Methods for determination of proline in plants. In: Sunkar R (ed) Plant stress tolerance. Humana Press, New York, pp 322–324
Alia P, Mohanty P, Matysik J (2001) Effect of proline on the production of singlet oxygen. Amino Acids 21:195–200
Alm V, Busso CS, Ergon A, Rudi H, Larsen A, Humphreys MW, Rognli OA (2011) QTL analyses and comparative genetic mapping of frost tolerance, winter survival and drought tolerance in meadow fescue (Festuca pratensis Huds.). Theor Appl Genet 123(3):369–382. doi:10.1007/s00122-011-1590-z
Atici O, Nalbantoglu B (2003) Antifreeze proteins in higher plants. Phytochemistry 64:1187–1196
Bajji M, Kinet JM, Lutts S (2001) The use of the electrolyte leakage method for assessing cell membrane stability as a water stress tolerance test in durum wheat. Plant Growth Regul 00:1–10
Bella J, Hindle KL, McEwan PA, Lovell SC (2008) The leucine-rich repeat structure. Cell Mol Life Sci 65:2307–2333
Bennett MD, Leitch IJ (2011) Nuclear DNA amounts in angiosperms: targets, trends and tomorrow. Ann Bot. doi:10.1093/aob/mcq258
Bocian A, Kosmala A, Rapacz M, Jurczyk B, Marczak Ł, Zwierzykowski Z (2011) Differences in leaf proteome response to cold acclimation between Lolium perenne plants with distinct levels of frost tolerance. J Plant Physiol 168:1271–1279
Bradbury PJ, Zhang Z, Kroon DE, Casstevens TM, Ramdoss Y, Buckler ES (2007) TASSEL: sotware for association mapping of complex traits in diverse samples. Bioinformatics 23:2633–2635
Brazauskas G, Lenk I, Pedersen MG, Studer B, Lübberstedt T (2011) Genetic variation, population structure, and linkage disequilibrium in European elite germplasm of perennial ryegrass. Plant Sci. doi:10.1016/j.plantsci.2011.06.013
Byrne S, Czaban A, Studer B, Panitz F, Bendixen C, Asp T (2013) Genome wide allele frequency fingerprints (GWAFFs) of populations via genotyping by sequencing. PLoS One. doi:10.1371/journal.pone.0057438
Campos PS, Quartin V, Ramalho JC, Nunes MA (2003) Electrolyte leakage and lipid degradation account for cold sensitivity in leaves of Coffea sp. Plants J Plant Physiol 160:283–292
Ciannamea S, Busscher-Lange J, de Folter S, Angenent GC, Immink RG (2006) Characterization of the vernalization response in Lolium perenne by a cDNA microarray approach. Plant Cell Physiol 47:481–492
Dinari A, Niazi A, Afsharifar AR, Ramezani A (2013) Identification of upregulated genes under cold stress in cold-tolerant chickpea using the cDNA-AFLP approach. PLoS One. doi:10.1371/journal.pone.0052757
Ebdon JS, Gagne RA, Manley RC (2002) Comparative cold tolerance in diverse turf quality genotypes of perennial ryegrass. Hortic Sci 37:826–830
Elshire RJ, Glaubitz JC, Sun Q, Poland JA, Kawamoto K, Buckler ES, Mitchell SE (2011) A robust, simple genotyping-by-sequencing (GBS) approach for high diversity species. PLoS One. doi:10.1371/journal.pone.0019379
Espevig T, Dacosta M, Hoffman L, Aamlid TS, Tronsmo AM, Clarke BB, Huang B (2011) Freezing tolerance and carbohydrate changes of two Agrostis species during cold acclimation. Crop Sci 51:1188–1197
Evanno G, Regnaut S, Goudet J (2005) Detecting the number of clusters of individuals using the software STRUCTURE: a simulation study. Mol Ecol 14:2611–2620
Forster JW, Cogan NOI, Dobrowolski MP, Francki MG, Spangenberg GC, Smith KF (2008) Functionally associated molecular genetic markers for temperate pasture plant improvement. In: Henry RJ (ed) Plant genotyping II: SNP technology. CABI Publishing, Cambridge, pp 154–186
Hardy OJ, Vekemans X (2002) SPAGeDi: a versatile computer program to analyse spatial genetic structure at the individual or population levels. Mol Ecol Notes 2:618–620
Hayat S, Hayat Q, Alyemeni MN, Wani AS, Pichtel J, Ahmad A (2012) Role of proline under changing environments: a review. Plant Signal Behav 7:1456–1466
Helft L, Reddy V, Chen X, Koller T, Federici L, Fernández-Recio J, Gupta R, Bent A (2011) LRR conservation mapping to predict functional sites within protein leucine-rich repeat domains. PLoS One. doi:10.1371/journal.pone.0021614
Hulke BS, Watkins E, Wyse D, Ehlke N (2007) Winter hardiness and turf quality of accessions of perennial ryegrass (Lolium perenne L.) from public collections. Crop Sci 47:1596–1602
Hulke BS, Watkins E, Wyse DL, Ehlke NJ (2008) Freezing tolerance of selected perennial ryegrass (Lolium perenne L.) accessions and its association with field winterhardiness and turf traits. Euphytica 163:131–141
Hulke BS, Bushman BS, Watkins E, Ehlke NJ (2012) Association of freezing tolerance to LpCBFIIIb and LpCBFIIIc gene polymorphism in perennial ryegrass accessions. Crop Sci. doi:10.2135/cropsci2011.09.0527
John UP, Polotnianka RM, Sivakumaran KA, Chew O, Mackin L, Kuiper MJ, Talbot JP, Nugent GD, Mautord J, Schrauf GE, Spangenberg GC (2009) Ice recrystallization inhibition proteins (IRIPs) and freeze tolerance in the cryophilic Antarctic hair grass Deschampsia antarctica E. Desv. Plant Cell Environ 32:336–348
Kaul S, Sharma SS, Mehta IK (2008) Free radical scavenging potential of l-proline: evidence from in vitro assays. Amino Acids 34:315–320
Lassner MW, Peterson P, Yoder JI (1989) Simultaneous amplification of multiple DNA fragments by polymerase chain reaction in the analysis of transgenic plants and their progeny. Plant Mol Biol Rep 7:116–128
Lee YP, Babakov A, Boer B, Zuther E, Hincha DK (2012) Comparison of freezing tolerance, compatible solutes and polyamines in geographically diverse collections of Thellungiella sp. and Arabidopsis thaliana accessions. BMC Plant Biol 12(1):131. doi:10.1186/1471-2229-12-131
Lissarre M, Ohta M, Sato A, Miura K (2010) Cold-responsive gene regulation during cold acclimation in plants. Plant Signal Behav 5:948–952
Meyer K, Keil M, Naldrett MJ (1999) A leucine-rich repeat protein of carrot that exhibits antifreeze activity. FEBS Lett 447:171–178
Middleton AJ, Brown AM, Davies PL, Walker VK (2009) Identification of the ice-binding face of a plant antifreeze protein. FEBS Lett. doi:10.1016/j.febslet.2009.01.035
Middleton AJ, Marshall CB, Faucher F, Bar-Dolev M, Braslavsky I, Campbell RL, Walker VK, Davies PL (2012) Antifreeze protein from freeze-tolerant grass has a beta-roll fold with an irregularly structured ice-binding site. J Mol Biol. doi:10.1016/j.jmb.2012.01.032
Muthukumaran J, Manivel P, Kannan M, Jeyakanthan J, Krishna R (2011) Advancement in computational analysis methods of plant antifreeze proteins (AFPs): an application towards classification and gene expression studies of leucine rich repeat (LRR) and ice-recrystallization inhibition domain (IRI) containing AFPs. J Comp Biol 3:65–82
Nekrošas S, Kemešytė V (2007) Breeding of ryegrass and Festulolium in Lithuania. Zemdirbyste–Agric 94(4):29–39
Pearce SR (2001) Plant freezing and damage. Ann Bot 87:417–424
Pritchard JK, Stephens M, Donnelly P (2000) Inference of population structure using multilocus genotype data. Genetics 155:945–959
Sandve SR, Rudi H, Asp T, Rognli OA (2008) Tracking the evolution of a cold stress associated gene family in cold tolerant grasses. BMC Evol Biol. doi:10.1186/1471-2148-8-245
Sandve SR, Kosmala A, Rudi H, Fjellheim S, Rapacz M, Yamada T, Rognli OA (2011) Molecular mechanisms underlying frost tolerance in perennial grasses adapted to cold climates. Plant Sci. doi:10.1016/j.plantsci.2010.07.011
Skøt L, Sanderson R, Thomas A, Skøt K, Thorogood D, Latypova G, Asp T, Armstead I (2011) Allelic variation in the perennial ryegrass FLOWERING LOCUS T gene is associated with changes in flowering time across a range of populations. Plant Physiol. doi:10.1104/pp.110.169870
Srinivas V, Balasubramanian D (1995) Proline is a protein-compatible hydrotrope. Langmuir 11:2830–2833
Statkevičiūtė G, Aleliūnas A, Kemešytė V, Pašakinskienė I, Brazauskas G (2014) AFLP analysis of genetic diversity in an association mapping panel of Lolium perenne L. In: Sokolović D, Huyghe C, Radović J (eds) Quantitative traits breeding for multifunctional grasslands and turf. Springer, Netherlands
Szabados L, Savouré A (2009) Proline: a multifunctional amino acid. Trends Plant Sci 15:89–97
Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: molecular Evolutionary Genetics Analysis Version 6.0. Mol Biol Evol 30:2725–2729
Thalhammer A, Hincha DK, Zuther E (2014) Measuring freezing tolerance: electrolyte leakage and chlorophyll fluorescence assays. Methods Mol Biol 1166:15–24
Tyler BF, Hayes JD, Davies WE (1987) Collection, characterization and utilization of genetic resources of temperate forage grass and clover. IBPGR, Rome, pp 1–65
Uemura M, Joseph RA, Steponkus PL (1995) Cold acclimation of Arabidopsis thaliana. Effect on plasma membrane lipid composition and freeze-induced lesions. Plant Physiol 109:15–30
Utz HF (1998) PLABSTAT: a computer program for the statistical analysis of plant breeding experiments. University of Hohenheim, Institute of Plant Breeding, Seed Science and Population Genetics, Stuttgart
Wang Z, Hopkins A, Mian R (2001) Forage and turf grass biotechnology. CRC Crit Rev Plant Sci 20:573–619
Warren RF, Henk A, Mowery P, Holub E, Innes RW (1998) A mutation within the leucine-rich repeat domain of the Arabidopsis disease resistance gene RPS5 partially suppresses multiple bacterial and downy mildew resistance genes. Plant Cell 10:1439–1452
Wilkins PW (1991) Breeding perennial ryegrass for agriculture. Euphytica 52:201–214
Xin Z, Browse J (2000) Cold comfort farm: the acclimation of plants to freezing temperatures. Plant Cell Environ 23:893–902
Xing Y, Frei U, Schejbel B, Asp T, Lübberstedt T (2007) Nucleotide diversity and linkage disequilibrium in 11 expressed resistance candidate genes in Lolium perenne. BMC Plant Biol. doi:10.1186/1471-2229-7-43
Xiong Y, Fei S, Arora R, Brummer EC, Baker RE, Jung G, Warnke SE (2007) Identification of quantitative trait loci controlling winter hardiness in an annual × perennial ryegrass interspecific hybrid population. Mol Breed. doi:10.1007/s11032-006-9050-1
Yoshiba Y, Kiyosue T, Nakashima K, Yamaguchi-Shinozaki K, Shinozaki K (1997) Regulation of levels of proline as an osmolyte in plants under water stress. Plant Cell Physiol 38:1095–1102
Yoshida S (1984) Studies on freezing injury of plant cells. I. Relation between thermotropic properties of isolated plasma membrane vesicles and freezing injury. Plant Physiol 75:38–42
Yu X, Bai G, Liu S, Luo N, Wang Y, Richmond SD, Pijut MP, Jackson AS, Yu J, Jiang Y (2013) Association of candidate genes with drought tolerance traits in diverse perennial ryegrass accessions. J Exp Bot 64:1537–1551
Zhang C, Fei SZ, Warnke S, Li L, Hannapel D (2009) Identification of genes associated with cold acclimation in perennial ryegrass. J Plant Physiol. doi:10.1016/j.jplph.2009.03.001
Zhang C, Fei SZ, Arora R, Hannapel DJ (2010) Ice recrystallization inhibition proteins of perennial ryegrass enhance freezing tolerance. Planta 232:155–164
Zhao MG, Chen L, Zhang LL, Zhang WH (2009) Nitric reductase-dependent nitric oxide production is involved in cold acclimation and freezing tolerance in Arabidopsis. Plant Physiol 151:755–767
Acknowledgments
This study was funded by the Research Council of Lithuania, Grant No. MIP-032/2012 (FUMAG). The authors acknowledge Vidmantas Feiza for his assistance in sequence analysis.
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Aleliūnas, A., Jonavičienė, K., Statkevičiūtė, G. et al. Association of single nucleotide polymorphisms in LpIRI1 gene with freezing tolerance traits in perennial ryegrass. Euphytica 204, 523–534 (2015). https://doi.org/10.1007/s10681-014-1330-y
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10681-014-1330-y