Sofija Bozinovic

High-throughput field phenotyping (HTFP) has become an important approach for improving the efficiency and objectivity of phenotypic evaluation in modern plant breeding. Within the PHENO_MaizE project the practical application of UAV-based RGB phenotyping in temperate maize breeding under field conditions is investigated. The project integrates repeated drone imaging, extraction of image-derived traits, and predictive modeling in order to evaluate the potential of digital phenotyping for the selection of superior maize genotypes. Experimental material includes maize inbred lines and their corresponding testcrosses evaluated across multiple environments in Serbia. UAV surveys conducted during the growing season will enable monitoring of temporal crop development and extraction of traits such as plant height, canopy cover, vegetation indices, and growth dynamics. The research within the project will also assess the potential of phenomic prediction models for estimating important agronomic traits, including grain yield, flowering time, and grain moisture at harvest. Special emphasis is placed on developing a practical, cost-effective, and scalable HTFP framework adapted to medium-sized breeding programs. The expected outcomes may support wider implementation of digital phenotyping and data-driven selection strategies in maize breeding.

Quality protein maize (QPM) has a high content of the essential amino acids lysine and tryptophan that are deficient in standard maize kernels. Maize Research Institute Zemun Polje has a breeding program aimed to create QPM, using the integrated conventional and molecular breeding approach. The aim of this study was biochemical evaluation of a QPM hybrid, obtained by crossing adapted QPM inbred lines (commercial lines converted to their QPM version). Relevant nutritional parameters, such as protein, starch, lipid and amino acid content were analyzed, and tryptophan to protein ratio (TPR) was determined. The tryptophan content (Trp) was significantly increased (p<0.05) in the QPM hybrid in comparison with standard maize (SM) hybrid. Also, the Trp was 0.075%, which corresponds to the QPM threshold value. The tryptophan to protein ratio, as an indicator of improved nutritional protein quality, was significantly higher (p<0.05) in the QPM hybrid. The starch and lipid content were both significantly higher (p<0.01 and p<0.05, respectively) compared to SM. Furthermore, a highly significant correlation (p<0.01) was identified between TPR and starch content, as well as a significant correlation (p<0.05) between TPR and lipid content, indicating the possibility of simultaneous selection and improvement of these traits in our breeding material. Biochemical evaluation has confirmed the improved nutritional value of this QPM hybrid, thus it was identified as a promising hybrid for use in the feed industry.

Marker assisted selection (MAS) is used in breeding to select progenies with the desired genes. MAS based gene pyramiding could facilitate in pyramiding of multiple genes for the simultaneous expression of more than one gene in a single genetic background. The main advantages of this strategy for germplasm improvement are elimination of extensive phenotyping, controlling the linkage drag and reducing the breeding duration. The aim of this study was to identify maize inbred lines that could serve as donors of opaque2 and crtRB1 favorable alleles. These genes can work together in the same genetic background to control the content of lysine, tryptophan, and beta-carotene. Three SSR markers, phi057, phi112 and umc1066, linked to the opaque2 gene, and crtRB1-3?TE, linked to the crtRB1, were employed for parental polymorphism screening. Recurrent parent (RP) was clearly distinguishable with crtRB1-specific marker from the donor parent (DP). The donors carried the favorable allele of crtRB1 (543 bp), while the recurrent parent exhibited the unfavorable crtRB1 allele (296 bp). Out of six DP lines, only one (DP1) was distinguishable with opaque2-specific markers from the RP line. The phi057 amplified 170 bp fragment in DP and 160 bp fragment in RP, while umc1066 amplified 150 bp fragment in DP and 160-170 bp fragment in RP. The phi112 showed a null fragment with DP and a band size around 170bp with RP. Therefore, DP1 was identified as the inbred line with potential to be used as a donor for pyramiding of opaque2 and crtRB1 genes, producing the breeding materials with high contents of lysine, tryptophan and beta- carotene, the major nutritional traits in maize.

Maize Research Institute Zemun Polje has a marker-assisted backcross breeding (MABB) program aimed to incorporate the strong allele of incompatibility gene Ga1 into parental components of the hybrids with specific traits (white kernel). The main objective of this research was marker-assisted development of the white maize hybrid's parental lines with complete cross-incompatibility to foreign pollens. Ga1-S-tightly-linked molecular markers successfully identified heterozygous plants with a percentage of approximately 50% after all three backcrossings. Regarding the background selection, 69% of progenies had recurrent parent's genome (RPG) above the theoretical value for BC3 generation. Also, a few individuals even had 99%, the value theoretically achieved in the BC6 generation, showing that MABB made a genetic gain in RPG recovery. Foreground selection in this generation aimed to identify homozygous dominant individuals. Out of 264 BC3 F2 plants, 70 (26.5%) were dominant homozygotes, which is in accordance with the expected Mendelian ratio. Progenies of the BC3 F2 homozygous dominant plants were planted alternatively with yellow-kernel maize of the same maturity to check cross-incompatibility. Unfortunately, none of the dominant progenies had a 100% white kernel, most likely due to the presence of modifier genes that increase/decrease the effectiveness of pollen exclusion or that, alternatively, modifiers are lost during the backcrossing. It could be expected that successfully employed functional markers (developed from the sequence polymorphisms present within a functional gene(s) associated with phenotypic trait variations) would outbalance the noted impediments and enhance MABB efficiency to transfer the desired gene(s) controlling simple or complex trait(s) into cultivated varieties.

Five direct and reciprocal early hybrids, five direct and reciprocal late hybrids and a total of their 12 parental inbred lines were used in order to test reciprocal effects among hybrids in early germination phase and seed quality of their parental inbred lines. Five hybrids were of early maturity FAO 300-400 group, and five of late maturity FAO 500-600 group. In the early group one line of Lancaster origin (L1) was in common, and in direct crosses used as male parent, while in reciprocal crosses as female parent. The same case was with late hybrids where another Lancaster line (L2) was in common. In 2015 and 2016 all the material was multiplied in hand pollination and after six months of storage three tests according to ISTA standards were performed: standard germination test (SGT), cold-test (CT) and vigor radicle test (RE). According to the Analysis of variance, a strong reciprocal effect was found among early hybrids for four out of five parameters in RE and SGT, while it was found only for RE in late hybrids. However, in CT none of the parameters had significant reciprocal effect. Two tested Lancaster inbreds were superior in the hybrids over non-Lancaster lines when used as maternal components, while as lines per se they did not display such superiority. Mid-parent heterosis (MPH) ranged from -62.32% to 211.11% in RE test, with an average of 44.26%; from -10.15% to 17.93% in SGT test, with an average of 6.27%, and from -5.31% to 4.49% in CT, with an average of 0.64%. Significant differences of MPH were found for nine pairs of reciprocal hybrids for RE, three pairs for SGT and seven pairs for CT. This shows that reciprocal differences in MPH were higher as the applied level of stress during early seed growth raised. All the results showed very complicated mode of inheritance of tested traits in the early developmental stages of young plants, including the phenomenon of heterosis which is expressed already at the germination stage.

With global climate change including unpredictable geographic and temporal weather patterns that lead to significant genotype ? environment interaction (GEI) the maize performance assessment would need to be complemented with stability analysis. The objectives of this study were: i) estimation of parametric and non-parametric stability indices for grain yield of late maturity maize hybrids, ii) assessing correlations and grouping of stability indices for grain yield of late maturity maize hybrids. The eight maize hybrids of FAO 700 maturity group (ZP1-ZP7, and check Ch) were evaluated for grain yield in the multi-environment trial including five locations in Serbia during 2020. and 2021. year. Stability analyses included twenty parametric and non-parametric indices, Principal Component Analysis (PCA) and Pearson?s correlations between stability indices. The sum of ranks for tested late maturity maize hybrids were in the interval from 37 (ZP6) to 146 (ZP4). The average rank was in the range from 1.8 (ZP6) to 7.3 (ZP4). The interval of variation for standard deviation of average ranks for tested maize hybrids was from 1.3 (ZP4, Ch) to 2 (ZP2) indicating satisfying accordance of utilized parametric and non-parametric stability measures. The highest average grain yield across all tested environments was observed for ZP1, which was second most stable hybrid. The most stable late maturity maize hybrid tested over ten environments was proven to be ZP6 with sum of ranks and average rank of 37 and 1.8, respectively, with standard deviation of average rank 1.5. PCA biplot showed two groups of parametric and non-parametric stability estimates: I group-coefficient of variation (CVi), Eberhart and Russel?s regression coefficient (bi), Perkins and Jinks?s regression coefficient (Bi), GEI variance component (?(i)), coefficient of determination (Ri2); II group-Eberhart and Russel?s deviation from regression (S2di) Wricke?s ecovalence (Wi2), Shukla?s stability variance (?i2)Perkins and Jink?s deviation from regression(?i2), superiority measure (Pi), the mean of the absolute rank differences of a genotype over all tested environments (Si(1))the variance among the genotype ranks over all tested environments (Si(2)), the sum of the absolute deviations for each genotype relative to the mean of ranks(Si(3)), the sum of squares of rank for each genotype relative to the mean of ranks (Si(6)), NPi(1), NPi(2), NPi(3), NPi(4)- Thennarasu non-parametric stability estimates, mean variance component (?i), Kang? rank-sum (KRi). Pearson?s correlation bring along the redundant stability indices and one can choose to use Wi2 or ?i2, bi or Bi, Wi2or ?i, ?i2 or ?i.

The Pannonian Plain, as the most productive region of Southeast Europe, has a long tradition of agronomic production as well as agronomic research and plant breeding. Many research institutions from the agri-food sector of this region have a significant impact on agriculture. Their well-developed and fruitful breeding programmes resulted in productive crop varieties highly adapted to the specific regional environmental conditions. Rapid climatic changes that occurred during the last decades led to even more investigations of complex interactions between plants and their environments and the creation of climate-smart and resilient crops. Plant phenotyping is an essential part of botanical, biological, agronomic, physiological, biochemical, genetic, and other omics approaches. Phenotyping tools and applied methods differ among these disciplines, but all of them are used to evaluate and measure complex traits related to growth, yield, quality, and adaptation to different environmental stresses (biotic and abiotic). During almost a century-long period of plant breeding in the Pannonian region, plant phenotyping methods have changed, from simple measurements in the field to modern plant phenotyping and high-throughput non-invasive and digital technologies. In this review, we present a short historical background and the most recent developments in the field of plant phenotyping, as well as the results accomplished so far in Croatia, Hungary, and Serbia. Current status and perspectives for further simultaneous regional development and modernization of plant phenotyping are also discussed.

Maize is a poor source of protein both for humans and monogastric animals due to the lack of essential amino acids, such as lysine and tryptophan. Naturally occurring opaque2 (o2) mutation increases content of these amino acids, but also confers an undesirable phenotype leading to low yields. Quality protein maize (QPM) is agronomically acceptable and nutritionally improved opaque2 maize obtained through conventional breeding. Marker assisted breeding program aimed at developing QPM genotypes for growing in temperate regions is being conducted at Maize Research Institute Zemun Polje (MRIZP). The results presented in this paper relate to foreground selection aimed to identify plants that attained homozygosity at o2 locus in BC2F2 generation in conversion of four MRIZP commercial inbred lines. Maize inbred line ZPL5 converted to its QPM counterpart and adapted to temperate climate was used as o2 donor to the four recurrent parents (RP). Foreground selection was carried out with gene-specific markers phi057 and umc1066, both segregating as per the expectation. The percentage of recessive homozygotes in BC2F2 generation was approximately 25% (24.6% in RP1, 23.3% in RP2, 25% in RP3 and 24.4% in RP4). After the self-pollination of selected recessive homozygotes, BC2F3 progenies were screened for phenotypic and biochemical characteristics to confirm their nutritional and agronomical superiority. The results of scoring endosperm modifications revealed over 95% of hard endosperm kernels. The average tryptophan content ranged from 0.070% in RP1 to 0.087% in RP3. Out of 39 derivations from four lines, 19 had tryptophan content above the QPM threshold (0.075%). A total of 16 derivations were chosen for their highest tryptophan content. Their quality index was increased by 2-46% relative to the recurrent parent. These line will serve as an important breeding material for developing QPM maize hybrids adapted to temperate regions.

The possibility of utilization of dried distillers' grains with solubles (DDGS) in animal feed was investigated. Samples of maize hybrids ZP 548 and ZP 655b, and DDGS obtained from hybrids ZP 548 and ZP 548c were used as components of the feed. The total protein content ranged from 12.42% to 31.18%, moisture from 5.49% to 9.55, and ash content from 1.85% to 4.37%. The contents of NDF, ADF, ADL, hemicellulose and cellulose fibers ranged from: 13.90% to 48.13%; 2.9% to 20.69%; 0.27% to 2.44%; 10.69% to 30.17%, 2.62% to 18.32%, respectively. In vitro, dry matter digestibility ranged from 55.20% to 89.76%. It was concluded that the samples of DDG obtained from red and yellow maize hybrids kernel are very suitable as components for the preparation of animal feed.

The aim of this study was to observe the properties of ten maize hybrids from Serbia including agronomic traits, lignocellulosic fibre composition and in vitro dry matter digestibility. Five yellow kernel dent hybrids and five analogue red kernel hybrids were used in this study to evaluate the agronomic traits, dry matter content of the whole plant, lignocellulosic fibre composition, ratios between different fibres, in vitro dry matter and NDF digestibility. Correlation coefficients between the investigated traits of the maize hybrids were assessed. Even though the results of our study showed variations regarding nutritional composition of the whole maize plant between the investigated maize hybrids, the hybrid and the differences in kernel colour (yellow or red) did not considerably affect the properties that influence quality of the maize hybrids for silage production. The highest IVDMD was determined in yellow kernel hybrid ZP 388, while the maximum NDFD was detected in the red kernel hybrid ZP 606red which also showed the lowest ADL/NDF and ADF/NDF ratios. The results indicate that all of the hybrids used in this study are good candidates for the production of high-quality silage for ruminant nutrition.