MORPHOLOGICAL PARAMETERS OF MAIZE ENDOSPERMAL MUTANTS SEEDLINGS AND EFFECT OF SEED STORAGE REGIMES ON THEM

Conclusions . Herbicide Pulsar 40 is the most suitable for soybean. Post-emergence application of Pulsar 40 in pea and chickpea crops without inoculation significantly reduced the 1000-seed weight. The greatest gain in the yield from seed inoculation with Rizobophyte was achieved in chickpea. There was no significant effect of the factors under investigation on the protein and fat contents in seeds. According to the reaction of morphological parameters of seedlings from the grains of maize endospermal mutants to the influence of the regimes that simulate a longterm storage – accelerated aging, freezing and combined regimes, the highest longevity is distinguished by ssp. indentata seeds, as well as carriers of the genes ae and su 1 , low durability – carriers of the genes wx and se , and the lowest – carriers of the gene sh 2 . Results and discussion. The decrease in the indices of all morphometric traits under influence of three factors was shown by carriers of the mutation sh 2 ♀SS386 and ♂SS389. Their F1 hybrid, as a rule, increased the indices and reduced only length of the seedling under the combined regime. Reduction of most indicators of morphometric traits under accelerated aging was observed in carriers wx : Binom, ♀VK69 and ♂VK64. Positive effect of accelerated aging and freezing was observed on the indices of all three morphological characters in ae carriers: AE800/AE392, ♀AE392, ♂ AE800. The excess of the hybrid over the parent lines in length of seedling and number of roots after accelerated aging and freezing manifested in the dentate hybrid Vympel, in the number and total length of roots – in carriers of sh 2 SS389 / SS386; in total roots length and seedling length – from the carrier su 1 Dmytryk. In general, the hybrids AE800/AE392 ( ae ) and SS389/SS386 ( sh 2 ) are more homeostatic than the parent lines. Conclusions. The highest longevity is characterized for the seeds of dentate maize and carriers of ae and su 1 , comparatively low longevity–carriers of wx and se , and the lowest – carriers of sh 2 .

It is believed that accelerated aging is a convenient and informative method that allows evaluating seed longevity upon natural aging and predicting it upon storage. This method consists in short-term incubation of seeds under high humidity and air temperature [13,14,15].
The international standards recommended the temperature -18±2 o C for long-term storage of seeds, which ensures the seed longevity [16]. This regime is also used in the National Repository of Plant Gene Pool Accessions of Ukraine which functions at the Plant Production Institute named after V.Ya. Yuriev of NAAS. In addition, previous studies [17] demonstrated a possibility of increasing the germinability of seeds which decreased it after accelerated aging, by incubation under -18±2 o C and thus reducing negative effects of aging. However, responses of seeds to these factors vary depending on the genotype [18]. Effects of accelerated aging and freezing of seeds of different maize endosperm mutations carriers have not been elucidated so far. This limits prospects for developing the most efficient storage technologies for this category of starting material for breeding.
In the previous article [19], we described the response of seeds of different corn endosperm mutants to two storage regimes in terms of the germination energy and germinability.
Our purpose was to determine effects of different regimes simulating long-term storage on morphological parameters of seedlings from the seeds of maize endosperm mutants.
All endosperm mutants were characterized by decreased content of starch compared to wild type corn, but among them, the wx mutation carriers had the highest average starch content and the sh 2 mutation carriers -the lowest one. The maize endosperm mutants were clearly divided into two groups by the average content of amylose in starch. Group I comprised the wx mutation carriers (almost amylose-free starch unlike wild type maize). Group II included the carriers of other mutations (ae, su 1 , se, and sh 2 ); they had higher amylose content in starch, and this effect was particularly pronounced in the ae mutation carriers compared to wild type dent maize.
The experiments were carried out with the seeds harvested in 2012-2014. 2012 and 2013 were characterized by increased average daily temperature with sufficient or excessive (at the end of vegetation) rainfall. Such conditions were favorable for grain filling, however, they delayed water evaporation from grain and full ripeness and favored formation of partly empty ears. 2013 was favorable for the spread of major diseases (common smut, stem rots, kernel rot) and corn pests (European corn borer and corn earworm). 2014 was dry, which contributed to fast ripening and low consumption of water by grain. Since the ranking of indices coincided in the three years, the article presents averaged data over the three years.
The experiments were conducted in three replicas, with 100 seeds for each. The longevity of seeds of maize lines and hybrids with a moisture content of 7-10% was studied in three models. Model 1 -«accelerated aging»: seeds of each form were acceleratedly aged by B.S. Likhachiov's method [15], which simulates natural aging during long-term storage. The seeds were placed in hermetically sealed foil bags and kept under 37°C for 30 days. Model 2 -«freezing»: exposure under -18 ± 2 °C for 1 month. This temperature is used in the National Repository of Ukraine for long-term storage of the plant gene pool accessions seeds. Model 3 -«combined mode»: after «accelerated aging», the seeds in the same sealed foil bag were exposed under room temperature for 4 hours and then placed in a freezer under -18 ± 2 ° C for 1-month storage.
Before the onset of each experiment and at each of its stages, the laboratory germination energy and germinability were evaluated in compliance with the State Standard 4138-2002 [21]. The seeds were sprouted in distilled water under 25 ± 1°C. Concurrently with evaluation of the geminability, morphological parameters were measured: sprout length, number and total length of primary roots [22]. The seeds of the same lines and hybrids (with the same moisture content) stored in paper bags under room temperature were taken as the control in the laboratory tests. The significance of differences between the means was estimated by t-test at probability level P = 95% [23].
The effects of accelerated aging, freezing and combined mode on the morphological parameters of seedlings were estimated by the I index [24]: % where: 1 X -experimental average indicator, 2 -control average indicator. A positive I means an increase in the parameter, and a negative I -a decrease.
Results and discussion. There were differences in the responses of dent and mutant maize seeds to accelerated aging, freezing and combined regimes manifested in the morphological parameters of seedlings (Figs. 1-6).  Control regime. The tested accessions significantly differed in the morphological parameters of seedlings.
Comparing the hybrids with their parent lines in the control, we noticed that no hybrid exceeded its parents by the root number. Two groups of accessions can be distinguished. In the group I the both parent lines had the roots numbers equal to their hybrid or one of the parent lines inferior to the hybrid. So, the hybrid Vympel (ssp. indentata) and its female line ♀HК26М had the identical root numbers of 3.4 ± 0.25 but its male line ♂Kh523ZM had a significantly (t> 3, P=95 %) smaller root number (1.6 ± 0.15). In the accessions with the wx endosperm mutation, the line ♂VК64 was almost at hybrid Binom's level (3.1 ± 0.25 and 3.7 ± 0.26, respectively), and the line ♀VК69 was inferior to them (2.5 ± 0.47). In accessions with the ae and su 1 endosperm mutations, hybrids also had almost the same roots number as a parent lines: ae -hybrid AE800/AE392 and lines ♀AE392, ♂АЕ800 (2.6 ± 0.10, 2.4 ± 0.06, 2.2 ± 0.08, respectively); Su1-hybrid Dmytryk and lines ♀MS401, ♂MS266 (1.7 ± 0.55, 1.2 ± 0.82, 1.2 ± 0.19, respectively).
In the group II, hybrids were inferior either to both parents or to one of them. Of the se endosperm mutation carriers, hybrid Snihova Koroleva and line ♂SE843 had the smallest root numbers (1.8 ± 0.21 and 1.2 ± 0.25, respectively) compared to the line ♀SE854 (2.6 ± 0.32), with significant difference (t> 3). Of the sh 2 endosperm mutation carriers, hybrid SS386/SS389 had the smallest number of roots (1.1 ± 0.21), and both parent lines ♀SS386, ♂SS389 had insignificantly exceeded it by the root number (1.7 ± 0.08 and 1.4 ± 0.30, respectively).
In general, the corn accessions with different endosperm structure under investigation were ranked in the control according to the three morphological parameters of seedlings as follows: high values of morphological parameters (the root number, their total length and seedling length) were intrinsic to hybrid Vympel and line ♀HK26M (dent type). High or intermediate number and total length of roots with intermediate or short seedlings were observed in hybrid Binom and its parent ♂VК64 (wx). Intermediate number and total length of roots with long seedlings were seen in the ae mutation carriers, hybrid AE800/AE392 and its parent lines ♀АЕ392 and ♂АЕ800. Low number of roots and short seedlings with intermediate total length of roots were intrinsic to lines ♀ МS401 and ♂МS266 (su 1 ). Long seedlings with various number and total length of roots is a common feature for parental lines -the se mutants ♀ SE854 and ♂SЕ843 and also sh 2 mutants ♀SS386 and ♂SS389. Hybrids with the su 1 and sh 2 genes were characterized by intermediate values of all the three morphological parameters.
There was a certain pattern between hybrids and parent lines. Hybrids AE800/AE392 (ae) and Dmytryk (su 1 ) significantly (t> 2) exceeded their parent lines by all the three morphological parameters; Vympel (dent type) and Binom (wx) significantly (t> 2, P=95%) exceeded their parent lines only by two parameters -the number of roots and their total length. Hybrid Snihova Koroleva (se) was inferior to its female line in terms of two morphological parameters (root number and length), and hybrid SS389/SS386 (sh2) -in terms of the three morphological features (seedling length, root number and length).
Accelerated aging. The effect of accelerated aging on seeds depended on the hybrid or line genotype. In general, stimulating effects were observed in 29 of 54 cases, i.e. in the majority; in 5 cases there was no response; depressing effects were recorded in 19 cases (Figs. 1-6).
The advantage of hybrids over parent lines after accelerated aging by all the three morphological features (t> 3, P=95%) was recorded in the sh 2 mutation carriers (SS389/SS386); by two features (the root number and their total length) -in the su 1 mutation carriers (Dmytryk). The hybrid was inferior to its parent lines by all the three indices in the ae mutation carriers (AE800/AE392). Thus, this hybrid was more homeostatic than its parent lines. Other hybrids were comparable with their parental lines or occupied an intermediate position between them by one, two or three indices.
The remaining hybrids and lines showed ambiguous responses to freezing: some indices were stimulated, others -inhibited or did not respond. The total number of indices in this category of accessions was 21, and stimulating effects were observed in 11 cases (most), no responsein 3 cases; and depressing effect -for 7 indices.
No hybrid showed exceeded its parent lines by all the three parameters after freezing. Hybrid Vympel (dent type) again exceeded both lines of its group by the total length of roots and the seedling length, hybrids Dmytryk (su 1 ) and SS389/SS386 (sh 2 ) -by the total length of roots (t> 3, p=95%). The remaining hybrids were comparable with both lines or occupied an intermediate position in terms of one, two or three morphological traits.
Stimulating effects (t> 2, p=95%) in the three indices were observed in the ae carriers: parent lines ♀ AE392 (I = 8.3 -32.5%) and ♂ AE800 (I = 4.5 -23.2%), while their hybrid responded to the combined regime by lowering all the three morphological parameters. The total length of roots was also increased in hybrid Snihova Koroleva (se, I = 15,2%).
In other accessions, the responses to the combined regime were various: of the total of 18 cases, stimulating effects were noticed in 6 cases; reduction -in 8 cases, no effect -in 3 cases.
Of the hybrids under investigation, Vympel (dent) exceeded its parents after the combined regime by all the three indices. The remaining hybrids were equal to the both parent, inferior to them or ranked intermediate by one -three morphological traits.
It was expected that hybrids would be characterized by smaller absolute values of the factor effect index compared to their parents, hence, be more homeostatic [25]. Concerning the response to accelerated aging, this occured in the ae mutation carriers -AE800 / AE392; after freezing, by the number and total length of roots -in AE800/AE392 (ae), Snihova Koroleva (se) and SS389/SS386 (sh 2 ) with seedlings lengths equal to those in the lines; by seed ling length -in Dmytryk (su 1 ); after the combined regime -in Vympel (dent corn), SS389/SS386 (sh 2 ) (in the latter, the seedling length was only equal to that in ŞS386); to a certain extent -in AE800/AE392 (ae), in which the total length of roots and seedling length were lower than those in parents or at least not exceeded them.
Summarizing the study results on the responses of the seedling morphological parameters to the factors that simulate the processes of storage of corn seeds with different endosperm structures, we established the following: All the morphological parameters were reduced under the influence of the three factors in the parent lines -the sh 2 mutation carriers, ♀SS386 and ♂SS389. Hybrid SS386/SS89, on the contrary, had either increased or unchanged indices. Reduction in the majority of indices under accelerated aging and combined regime was shown for all the wx mutation carriers: hybrid Binom and its parents ♀ VK69 and ♂VK64. Positive effects of all the three factors on almost all morphological parameters were noticed in the ae mutation carriers: parent lines ♀АЕ392 and ♂АЕ800. Positive effects of accelerated aging and freezing were observed in these lines and their hybrid AE800/AE392. Hybrids exceeded parent lines by the majority of morphological parameters in dent accessions (Vympel); by the total length of roots -in the su 1 mutation (Dmytryk) and sh 2 mutation (SS389/SS386) carriers.
The correlation analysis showed that the number of roots, their total length and length of seedlings positively correlated with the germinability in the accelerated aging, freezing and combined regime experiments: r = 0.47-0.64. Relatively high correlations were found between the germinability and the effects on the seedling length after accelerated aging and freezing: r = 0.72 and 0.75, respectively; the coefficient was somewhat lower after the combined regime: r = 0.64. The correlation coefficients from sub-moderate to moderate, also positive, were noticed between the germinability and the number of roots and their total length in all the three experimental variants: r = 0.27 -0.63. Consequently, the stress factors under investigation had similar effects on the germinability and morphological characteristics of corn seedlings, with the strongest effect on the seedling length. At the same time, the correlations between the initial (control) germinability and morphological parameters of seedlings after influence of the three studied factors were weak: r did not exceed 0.35. Thus, the initial germinability cannot predict the development of seedlings after influence of these stress factors.

Conclusions.
Summarizing the results of studying the response of the seedling morphological characteristics to the factors that simulate the processes of storage of corn seeds with different endosperm structures, we established the following: The three factors suppressed all the morphological characteristics in the parent lines -sh 2 mutation: ♀SS386 and ♂SS389. At the same time, hybrids, on the contrary, more often had increased indices, and only in one case a hybrid reduced them (the seedling length under the combined regime). Decrease in the majority of morphological characteristics under accelerated aging occurred in the wx mutation carriers: hybrid Binom and its parental lines ♀VK69 and ♂VK64. Positive effects of two stress factors (accelerated aging and freezing) were noted for all the morphological characteristics in the ae mutation carriers: hybrid AE800/AE392 and parent lines ♀АЕ392 and ♂АЕ800. The remaining hybrids and lines showed ambiguous responses to the three stress factors: some indices were stimulated, others -inhibited or did not respond.
After accelerated aging and freezing, hybrids to various degrees exceeded their parent lines by two morphological characteristics (seedling length and root number) in dent accessions (Vympel); by number and total length of roots -in the sh 2 mutation carriers (SS389/SS386); and by the total length of roots and seedling length -in the su 1 carriers (Dmytryk).

MORPHOLOGICAL PARAMETERS OF MAIZE ENDOSPERMAL MUTANTS SEEDLINGS AND EFFECT OF SEED STORAGE REGIMES ON THEM
Shyianova T.P., Tymchuk S.M., Boguslavskyi RL. Plant Production Institute nd. a V.Ya. Yuriev of NAAS, Ukraine Endospermal mutants of maize are a source of valuable raw materials for various industries and directions of consumption. Therefore, problem of their seeds longevity is relevant for storage in genebanks, breeding and seed-growing establishments. The aim and tasks of the study was to determine effect of different storage regimes on morphometric characteristics of seedlings from seeds of maize endosperm mutants. Material and methods. Seeds of six simple hybrids F1 and 12 their parental lines -carriers of endospermal mutations: wx, ae, su 1 , se, sh 2 , and dentate type are used as the material. The seeds were affected by accelerated aging using the method of B.S. Likhachev (1978), freezing during a month and combined regime -accelerated aging + freezing.
Results and discussion. The decrease in the indices of all morphometric traits under influence of three factors was shown by carriers of the mutation sh 2 ♀SS386 and ♂SS389. Their F1 hybrid, as a rule, increased the indices and reduced only length of the seedling under the combined regime. Reduction of most indicators of morphometric traits under accelerated aging was observed in carriers wx: Binom, ♀VK69 and ♂VK64. Positive effect of accelerated aging and freezing was observed on the indices of all three morphological characters in ae carriers: AE800/AE392, ♀AE392, ♂ AE800. The excess of the hybrid over the parent lines in length of seedling and number of roots after accelerated aging and freezing manifested in the dentate hybrid Vympel, in the number and total length of roots -in carriers of sh 2 SS389 / SS386; in total roots length and seedling length -from the carrier su 1 Dmytryk. In general, the hybrids AE800/AE392 (ae) and SS389/SS386 (sh 2 ) are more homeostatic than the parent lines. Conclusions. The highest longevity is characterized for the seeds of dentate maize and carriers of ae and su 1 , comparatively low longevity-carriers of wx and se, and the lowest -carriers of sh 2 .