Physiological and genetic features of the rate of development of modern varieties of soft wheat (Triticum aestivum L.)

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Selection and Genetic Institute -- National Center of Seed Science and Variety Studies

Odesa National Agrarian University

Physiological and genetic features of the rate of development of modern varieties of soft wheat (Triticum aestivum L.)

Fayt V., Hubich О., Nahuliak O., Balashova І., Fedorova V., Zelenina H.

Ovidiopolska Doroha, Odesa, 65036, Ukraine,

13 Panteleimonivska Str., Odesa, 65012, Ukraine

Goal. Assessment of levels of vernalization demand, photoperiodic sensitivity, frost resistance, and identification of alleles of Vrn1 and Ppd1 genes of two-handed varieties of soft wheat. Methods. Field: growing plants in the conditions of shortened and extended days of phytotron and vegetation site; hybridological analysis by photoperiodic sensitivity genes (Ppd1) and type of development (Vrn1); analysis of variance and correlation; criterion c2; multiplex STS-PCR with specific primers to the PpdD1 gene. Results. Phenotypic differences of the studied varieties in response to vernalization and sensitivity to the photoperiod were revealed. 5 groups of varieties with different Vrn1 and 2 groups of varieties with different Ppd1 genotypes were identified. Winter and frost resistance of two-handed varieties in the tillering and seedling phase, as well as the reaction of two-handed varieties for the duration of the period before earing to winter and spring sowing dates, were assessed. Conclusions. Varieties Demir 2000, Shestopalivka are winter varieties with low sensitivity to the photoperiod, others are typically spring with weak (Solomiia, Pallada, Afina, Yara, L897Я23) or strong (Lastivka, Khutorianka, Zimoiarka) sensitivity to the photoperiod. Weak reaction to the photoperiod of Afina, Pallada, Solomiia, Shestopalivka, Yara, Demir 2000, L897Я23 varieties is caused by the PpdD1a gene. Varieties Lastivka, Khutorianka, Zimoiarka are carriers of only recessive alleles of 3 genes of the orthological series Ppd1. The spring type of development of Zimoiarka and Khutorianka varieties is caused by 2 genes VrnA1a and VrnB1a, Afina, Lastivka, L897Я23 -- by genome VrnD1a, Solomiia -- by VrnA1a, Pallada and Yara -- by VrnB1a. The presence in the genotype of varieties at once of two genes -- VrnA1a and VrnB1a, or only one -- VrnA1a, and in some cases -- only VrnD1a, contributes to a significant reduction in frost resistance of seedlings and winter hardiness of two-handed plants. Winter and spring sowing dates cause the shift of earing time to much later calendar dates compared to sowing in autumn, which can negatively affect the formation of the yield of two-handed varieties.

Key words: Vrn1 and Ppd1 genes, vernalization, photoperiod, earing, winter, frost resistance.

Introduction

The response of plants to low positive temperatures (vernalization) and the duration of lighting (photoperiodism) are the basic mechanisms of the regulation process of wheat ontogenesis [1]. The response to vernalization in the initial ontogenesis period for the subsequent passage to generative development is the main feature that allows distinguishing typically winter wheat genotypes from typically spring. Winter wheat has a significant need for vernalization, while the spring may be non-sensitive or partially sensitive to vernalization. Both spring and winter genotypes can be sensitive or non-sensitive to the photoperiod. Qualitative differences in type of development are controlled by the genes of the orthologous series Vrn-1. The winter type of development is caused by the presence in the genotype only recessive alleles of these genes: Vrn-A1b, Vrn-B1b, Vrn-D1b. The presence in the genotype any dominant allele Vrn-A1a, Vrn-B1a, Vrn-D1a determines the spring type of development [2, 3]. The differences in photoperiodic sensitivity are due to the action of three Ppd-1 genes located on the chromosomes of the second homeological group: Ppd-A1, Ppd-B1, Ppd-D1. The dominant alleles of these genes contribute to a decrease in sensitivity to the photoperiod, and a significant response to the photoperiod is inherent in genotypes with recessive alleles of all three genes [4, 5]. Transitional, alternative, facultative spring or facultative winter varieties are the genotypes that can develop and pass on to the generative development at both spring and autumn sowing. Alternative wheat is grown mostly in areas with mild winters. In the south of Ukraine, alternative varieties are suitable for late autumn and winter crops, in years with prolonged dry autumn [6]. The peculiarity of alternative varieties is a high sensitivity to shortening the day length and a spring type of development, that is, a inexpressive reaction to the vernalization [7, 8]. The significant delay in the development of alternative varieties in the autumn is due to the interaction of the Vrn-1 and Ppd-1 genes, in particular the dominant Vrn-B1a gene with recessive alleles Ppd-A1b, Ppd-B1b and Ppd-D1b [7]. At the same time, some authors [8] note the possibility of controlling the spring type of development of alternative varieties by dominant alleles of genes Vrn-D1 or even Vrn-A1. Genotypes with dominant Vrn-A1a gene that do not respond to the photoperiod are typically spring varieties [9]. All other combinations of Vrn-1 and Ppd-1 genes effect on differences in spiking times [10] and resistance to frost [11]. Such genotypes can only be assigned to alternative in mild winters. However, hardening in both extended and shortened day duration provided a higher frost resistance of alternative varieties with the dominant Vrn-B1a gene [8, 12].

In numerous recent scientific and promotional publications on alternative wheat in Ukraine, unfortunately, there is no complete research on the physiology of development (ontogenesis) and the genetics of alternative genotypes [13, 14, 15]. This often does not determine the difference between true alternative and short-term winter varieties or spring cold-resistant genotypes.

The purpose of the study was to assess the levels of vernalization needs, photoperiodic sensitivity, frost resistance, and identification of bread wheat by alleles of Vrn-1 and Ppd-1 genes.

Materials and methods of researches. Varieties Afina, Lastochka, Pallada, Yarka, L897Я23 (Krasnodar Scientific Research Institute of Agriculture named after P.P. Luk'ianenko, Russia), Khutorianka, Zymoiarka (Institute of Plant Physiology and Genetics National Academy of Sciences of Ukraine, Kyiv), Solomiia (Kherson State Agrarian Univercity, Kherson), Shestopolivka (farm "BOR") were used as starting breeding material which are described by their authors as alternative [16-19]. Also were used variety Demir 2000 (which, according to the description of the Center for Genetic Resources of Ukraine, is alternative) and F2 populations from the diallelic crossings of this varieties and crossing with almost isogenic monogeneously dominant by the Vrn-1 genes of the Mironovskaya 808 or Skorospelka 3b populations. In various experiments, the winter variety Borvii of the PBGI-NCSCI breeding and alternative almost isogenic breeding line of the Mironovskaya 808 variety according to the Vrn-B1a gene (hereinafter Mironovskaya 808 Vrn-B1a) were used as a controls.

Seeds of varieties and F2 populations from diallelic crossing were germinated at room temperature. Five-day seedlings were vernalized in the special chamber at + 2 °C and for a duration of day 12 hours: for varieties - 40, 30, 20, 10 days; for F2 populations - 40 days. After the end of the vernalization, the seedlings were planted under artificial conditions in light phytotron chambers, ten plants in 5-liter vessels and one part of each variety was grown under a 16- hour prolonged day and the rest and F2 populations - under a 12-hour shortened day. At the same time, not vernalizated five-day seedlings were planted under prolonged and shortened days. The reaction to the vernalization of a particular sample was determined by comparing the average date of its earing in two adjacent variants of the previous vernalization. The difference between the average duration of the period before earning of the variety after the vernalization under growing on the shortened and elongated day characterized its level of photoperiodic sensitivity.

Genetic analysis of half-diallelic F2 hybrids by photoperiodic sensitivity was performed by this method [20]. The distribution of F2 populations into the phenotypic classes of plants, which sooner and later earned under shortened day, was carried out according to the date of earning of the first plant of the Mironovskaia line 808 Vrn-B1a. Multiplex PCR with gene-specific primers was used to determine alleles of Ppd-D1a and Ppd-D1b [21].

Hybridological analysis by type of development and distribution of F2 populations into the phenotypic classes of spring and winter plants was performed by the this method [22], with the only difference being that the plants were grown in 5-liter pots (ten plants per pot) at the growing area.

Frost resistance was assessed by freezing seedlings at -12⁰C and using the “beams” method at -16⁰C [23] in the freezing chamber. Winter hardiness was assessed in the field by accounting for plants in autumn and in spring season. Seeds of alternative varieties Demir 2000, Afina, Zymoiarka, L-897Y23, Lastivka, Pallada, Solomiia, Khutorianka, Shestopolivka, Yara and control samples winter variety Borvii and alternative line Mironovskaya 808 Vrn-B1a were sowned in autumn 2012, 2014 and 2015 (October 15, 9 & 20, respectively), as well as in different calendar terms during the winter and spring of 2013 (19.02, 05.03, 19.03 and 02.04) and 2014 (24.12.2014 and 25.02.2015) by manual planting on the experimental area of the General and Molecular Genetics Department of of PBGI - NCSCI on two row sections with length of 1 m by 20 grains per row with plant feeding area of 30х5 cm². Repeat the experiment - two to three times.

Dates of spiking of individual plants were marked in the phytotron and at the vegetation area during the growing season for determine the period before earning. In the field, the date of spiking was marked visually in the presence of 75% of the spiked plants in the area.

Statistical processing of the obtained results was carried out according to conventional methods of variance and correlation analysis, criterion %² [24].

Research results

Comparison of the period duration before the spiking of alternative varieties plants after vernalization of different duration (term) under prolonged (PD) and shortened (SD) day of the climatic chambers of the phytotron indicated a significant influence of the genotype on the specified trait (Table 1). If as a criterion for evaluating the reaction to vernalization use the fact of earning or not earning a specific genotype after pre-vernalization of a certain duration, then the studied varieties can be divided into two groups. Varieties of the first group Afina, Zymoiarka, Lastivka, Pallada, Solomiia, Khutorianka, Yara, L897Я23 and control alternative line Mironovskaya 808 Vrn-B1a spiked by 41.6 days (Afina) - 89.9 days (Pallada) in conditions of PD regardless of the previous vernalization of different duration (40-10 days) and even in its absence. In the conditions of SD, the earning of the varieties Afina, Zymoiarka, Lastivka, Pallada, Solomiia, Khutoryanka, Yara, L897Y23 were noted at 43.1 days (Afina) - 97.5 days (Lastivka) both in the variant without vernalization, and after 10-40 days of the previous vernalization. However, in the absence, as well as in the vernalization of 10 and 20 days, the spiking of plants of the almost isogenic line Mironovskaya 808 Vrn- B1a (alternative genotype, control) was not observed in the conditions of SD. The gradual reduction of the vernalization duration to 30, 20, 10 days and to its complete absence contributed to the increase in the duration of the spiking period by 2.1 days (Solomiia) - 37.5 (Pallada) days in the variance without vernalization compared to the 40-day vernalization in the conditions PD and 8.3 days (Solomiia) - 41.3 (Yara) days in the conditions of SD. However, these genotypes differ in response to vernalization. Thus, in the artificial conditions of the phytotron chambers, the variety Solomiia almost did not respond to the vernalization by the acceleration of development (shortening the duration of the period to the earning). Zymoyarka and Khutoryanka varieties are smaller, and Lastivka - to a greater accelerated the development after 10 days of vernalization. The Afina and Yara varieties and the L897Я23 line responded significantly even to the 20-day vernalization. Of all the genotypes, the Pallada variety and the Mironovskaya 808 Vrn-B1a control line, which responded for 30 days of vernalization with the acceleration of the ear, are slightly different.

The varieties of the second group - Shestopalivka and Demir 2000 - responded to the vernalization similar to the control winter variety Borvii. The need for vernalization of the Borvii was 40 in the conditions of PD, and 30 days - of SD . Earning of varieties Shestopalivka and Demir 2000 were noted both in the PD and in the SD only after 30 days of vernalization. At the same time, in both cases, the 40-day vernalization significantly accelerated the spiking of these varieties by 16.9 - 21.3 and 18.4 - 19.7 days, respectively.

Table 1 The duration of the period before spiking of alternative varieties in the conditions of prolonged (PD) and shortened (SD) days after temporal vernalization, days