1 Introduction

The Triticeae tribe, a group of cereal grasses, has been a cornerstone of human agriculture since its inception. This tribe includes some of the most vital crops such as wheat, barley, and rye, which have been the primary food sources in temperate regions for millennia (Mascher et al., 2017). The genetic diversity within Triticeae, encompassing cultivated, wild, and weedy taxa, offers a rich reservoir of traits that can be harnessed for crop improvement and ensuring global food security (Lu and Ellstrand, 2014).

Triticeae species are indispensable in agriculture due to their significant contributions to food production. Wheat (Triticum spp.), for instance, is a staple food crop globally, providing essential nutrients and calories to a large portion of the world's population (Hyun et al., 2020). Barley (Hordeum vulgare L.) is another critical crop, known for its adaptability to diverse environmental conditions and its use in food, feed, and brewing industries (Mascher et al., 2017). Rye (Secale cereale L.), although less widespread, is crucial in Central and Eastern Europe for its resilience to poor soils and harsh climates (Bauer et al., 2017). The genetic improvement of these crops is vital for enhancing yield, disease resistance, and stress tolerance, thereby supporting sustainable agriculture and food security.

Molecular markers are powerful tools in genetic research, enabling the identification and mapping of genes associated with desirable traits. Techniques such as Genotyping-by-Sequencing (GBS) and Single Nucleotide Polymorphism (SNP) markers have revolutionized the study of genetic diversity and phylogeny within Triticeae (Hyun et al., 2020). The development of databases like TriMEDB, which integrates transcribed markers, facilitates advanced genetic studies, including quantitative trait loci (QTL) analysis and population genomics (Mochida et al., 2008). These markers are essential for marker-assisted selection (MAS), accelerating the breeding process by allowing precise tracking of genetic traits.

This study explores the application of Structural Equation Modeling (SEM) and Start Codon Targeted (SCoT) markers in the genetic improvement of Triticeae crops. By assessing the current state of genetic resources and molecular marker technologies within the Triticeae tribe, evaluating the effectiveness of SEM and SCoT markers in identifying and mapping key genetic traits, and discussing the potential applications of these markers in breeding programs aimed at enhancing crop yield, disease resistance, and stress tolerance, this research provides a comprehensive understanding of the role of SEM and SCoT markers in Triticeae genetic improvement. It emphasizes their potential to promote sustainable agricultural practices and contribute to food security.

2 Methodological Advances in SEM and SCoT Markers

2.1 Advances in scanning electron microscopy (SEM)

Scanning Electron Microscopy (SEM) has seen significant advancements over the past decades, greatly enhancing its application in various scientific fields, including plant genetics. Recent developments in SEM technology have focused on improving resolution and imaging capabilities (Lenthe et al., 2018). Innovations such as field emission electron sources, which provide higher brightness and resolution, have enabled researchers to observe plant structures at the nanoscale. Additionally, advancements in detector technologies, including the use of backscattered electron detectors, have improved the contrast and detail of images, allowing for more precise visualization of cellular and subcellular structures (Pease, 2008).

Moreover, new preparation techniques have been developed to preserve the ultrastructure of biological samples, including cryo-fixation and freeze-substitution methods. These techniques are essential for maintaining the integrity of delicate plant tissues during the imaging process (Bobik et al., 2014). The combination of high-resolution imaging and advanced sample preparation has expanded the utility of SEM in studying plant genetics, enabling detailed analysis of Triticeae species' morphological and anatomical features that are critical for understanding genetic variations and improvements (Souza and Attias, 2018).

2.2 Development and utilization of start codon targeted (SCoT) markers

Start Codon Targeted (SCoT) markers are a relatively recent addition to the molecular toolkit for plant genetics and breeding. These markers are designed to target the conserved regions flanking the start codon (ATG) of plant genes, making them useful for detecting genetic polymorphisms (Amom et al., 2020). SCoT markers are advantageous because they are simple, cost-effective, and require no prior sequence information, making them accessible for a wide range of genetic studies.

The development of SCoT markers has significantly advanced the genetic analysis of Triticeae species. They have been successfully used in various applications, including assessing genetic diversity, phylogenetic studies, and marker-assisted selection. For instance, SCoT markers have been employed to identify genetic variations and construct genetic linkage maps in wheat, facilitating the discovery of genes associated with important agronomic traits such as disease resistance and yield improvement (Collard and Mackill, 2008).

2.3 Comparative analysis of SEM and SCoT markers with other molecular markers

When comparing SEM and SCoT markers with other molecular markers, such as Simple Sequence Repeats (SSRs) and Single Nucleotide Polymorphisms (SNPs), several distinct advantages and limitations emerge. SEM provides detailed morphological and anatomical insights at the microscopic level, which are crucial for understanding the physical manifestations of genetic traits (Nadeem et al., 2018). However, SEM does not directly provide information on genetic polymorphisms or molecular variations.

On the other hand, SCoT markers, along with SSRs and SNPs, offer precise genetic information that can be used for constructing genetic linkage maps and identifying quantitative trait loci (QTLs). While SSRs are highly polymorphic and co-dominant, making them ideal for genetic diversity studies, SNPs provide the highest resolution of genetic variation and are abundant throughout the genome. SCoT markers, although not as high-resolution as SNPs, are valuable for their simplicity and broad applicability without requiring extensive genomic resources (Collard and Mackill, 2008).

3 Applications of SEM and SCoT Markers in Triticeae

3.1 Use of SEM in studying morphological traits

Scanning Electron Microscopy (SEM) has proven to be a valuable tool in the study of morphological traits in Triticeae. SEM allows for high-resolution imaging of plant structures, providing detailed insights into the surface morphology of grains and other plant parts. For instance, a study on Northern African Triticum aestivum L. cultivars utilized SEM to examine the grain surface sculpture, revealing distinct patterns on both the dorsal and ventral surfaces. This detailed morphological characterization facilitated the discrimination and identification of different cultivars, highlighting the utility of SEM in taxonomic studies and genetic diversity assessments (Mohamed et al., 2017).

Moreover, SEM has been instrumental in understanding the morphological variations that occur due to environmental factors or genetic differences. By providing a clear and magnified view of plant structures, SEM helps in identifying subtle morphological traits that are not easily discernible through conventional microscopy. This capability is particularly useful in breeding programs where precise morphological characterization is essential for selecting desirable traits (Mohamed et al., 2017).

3.2 Genetic diversity studies using SCoT markers

Start Codon Targeted (SCoT) markers have emerged as a powerful tool for assessing genetic diversity in Triticeae. These markers are based on the short conserved region flanking the ATG start codon in plant genes, making them highly reproducible and informative. In a study involving hexaploid wheat (Triticum aestivum L.) and two Aegilops species, SCoT markers were used to evaluate molecular variability among 91 samples. The results demonstrated high polymorphism and effective grouping of samples based on their genomic constitutions, underscoring the effectiveness of SCoT markers in genetic diversity studies (Figure 1) (Ghobadi et al., 2021).

Ghobadi et al. (2021) conducted a population structure analysis on 91 germplasm accessions of wheat (Triticum) and goatgrass (Aegilops) based on SCoT (a) and CBDP (b) markers. The results revealed three major sub-populations (Sub-Pop1, Sub-Pop2, Sub-Pop3), with colors representing different genetic components. The SCoT markers demonstrated clear population structure delineation, highlighting their effectiveness in studying plant genetic diversity. Compared to other markers, SCoT markers not only excelled in revealing genetic differences within populations but also accurately distinguished genetic relationships between different sub-populations, providing a more comprehensive understanding of the genetic background. This further validates the reliability and utility of SCoT markers for genetic diversity analysis in complex genomic contexts.

SCoT markers have also been used to assess genetic diversity in barley (Hordeum vulgare L.). In a study involving ten barley lines, SCoT markers revealed significant polymorphism and identified unique markers associated with specific traits. This information is crucial for breeding programs aimed at improving specific traits such as grain filling period and yield. The high resolving power and polymorphic information content of SCoT markers make them a valuable tool for genetic diversity studies and marker-assisted selection in Triticeae (Habiba et al., 2021).

3.3 Integration of SEM and SCoT markers for comprehensive genetic analysis

The integration of SEM and SCoT markers offers a comprehensive approach to genetic analysis in Triticeae. By combining detailed morphological characterization with molecular marker analysis, researchers can gain a holistic understanding of genetic diversity and relationships among different cultivars (Ibrahim et al., 2017). For example, a study on Northern African Triticum aestivum L. cultivars utilized both SEM and SCoT markers to assess taxonomic and genetic diversity. The combined analysis provided complementary insights, with SEM revealing morphological traits and SCoT markers elucidating genetic relationships. This integrated approach proved efficient in characterizing the genetic diversity of the studied cultivars (Mohamed et al., 2017).

Such integration is particularly beneficial in breeding programs where both morphological traits and genetic markers are important for selecting superior genotypes. By using SEM to identify desirable morphological traits and SCoT markers to assess genetic diversity, breeders can make informed decisions to enhance crop improvement efforts. This comprehensive approach ensures that both phenotypic and genotypic variations are considered, leading to more effective and targeted breeding strategies (Mohamed et al., 2017).

4 Advantages of SEM and SCoT Markers

4.1 Benefits of SEM in genetic studies

Scanning Electron Microscopy (SEM) offers several advantages in genetic studies, particularly in the detailed visualization of cellular and subcellular structures. SEM provides high-resolution images that allow researchers to observe the intricate architecture of chromosomes and other cellular components. For instance, the combination of Focused Ion Beam (FIB) milling with SEM has enabled the high-resolution three-dimensional reconstruction of metaphase barley chromosomes, revealing the chromatin packaging and the distribution of histone variants within the chromosome interior (Schroeder-Reiter et al., 2009). This level of detail is crucial for understanding the spatial organization of genetic material and its functional implications.

Moreover, SEM's ability to visualize biological samples without heavy metal staining enhances its utility in genetic studies. Traditional methods often require staining, which can obscure fine details and alter the sample's native state. However, advancements in SEM techniques, such as the use of cryo-FIB-SEM, allow for the imaging of fully hydrated, close-to-native cells without staining, preserving the biological material's integrity (Spehner et al., 2020). This capability is particularly beneficial for studying the native state of genetic material and cellular structures, providing insights that are closer to their natural conditions.

4.2 Advantages of SCoT markers in genetic diversity studies

Start Codon Targeted (SCoT) markers are highly effective in assessing genetic diversity due to their ability to target functional regions of the genome. SCoT markers are designed to amplify regions flanking the start codon (ATG) of genes, which are generally conserved and functionally significant (Feng et al., 2015). This targeting increases the likelihood of detecting polymorphisms that are directly related to phenotypic traits, making SCoT markers particularly useful for genetic diversity studies in Triticeae and other plant species.

One of the primary advantages of SCoT markers is their reproducibility and high polymorphism rates (Igwe et al., 2021). These markers generate clear and distinct banding patterns, which facilitate the accurate assessment of genetic variation among different genotypes. This high level of polymorphism is essential for distinguishing closely related individuals and for identifying unique genetic traits that can be harnessed for breeding programs. Additionally, SCoT markers do not require prior sequence information, making them a cost-effective and versatile tool for genetic diversity studies across a wide range of species (Elshire et al., 2011).

4.3 Synergistic benefits of using SEM and SCoT markers together

The combined use of SEM and SCoT markers offers synergistic benefits that enhance genetic improvement efforts in Triticeae. SEM provides detailed morphological and structural insights at the cellular and subcellular levels, which can be correlated with genetic data obtained from SCoT markers. For example, SEM can reveal the physical manifestations of genetic variations, such as changes in chromosome architecture or cellular morphology, which can then be linked to specific genetic markers identified by SCoT analysis (Schroeder-Reiter et al., 2009; Kuipers and Giepmans, 2015). This integrated approach allows for a more comprehensive understanding of the relationship between genotype and phenotype.

Furthermore, the high-resolution imaging capabilities of SEM can be used to validate and complement the genetic data obtained from SCoT markers. By visualizing the cellular and chromosomal structures associated with specific genetic traits, researchers can confirm the functional significance of the polymorphisms detected by SCoT markers. This validation is crucial for ensuring the accuracy and reliability of genetic improvement programs. Additionally, the ability to observe the native state of biological samples using advanced SEM techniques, such as cryo-FIB-SEM, enhances the relevance of the findings to real-world conditions (Spehner et al., 2020; Uryu et al., 2023). This holistic approach, combining the strengths of both SEM and SCoT markers, provides a powerful framework for advancing genetic research and breeding efforts in Triticeae.

5 Case Studies in Triticeae

5.1 Genetic analysis of Northern African Triticum aestivum cultivars

SEM and SCoT markers revealed significant taxonomic and genetic insights into Northern African Triticum aestivum cultivars. In a study by Mohamed et al. (2017), fourteen T. aestivum cultivars from seven Northern African countries were analyzed using high-resolution SEM imaging and SCoT polymorphism analysis. The SEM analysis identified distinct grain surface sculptures, while SCoT markers characterized the genetic diversity among the cultivars. The combined use of SEM and SCoT markers provided a comprehensive understanding of the taxonomic and genetic diversity, suggesting that these cultivars could serve as valuable genetic resources for future breeding programs (Mohamed et al., 2017).

5.2 Analysis of genetic variability in Aegilops and Triticum species

SCoT markers have been instrumental in assessing genetic relationships among progenitors of bread wheat. Pour-Aboughadareh et al. (2018) utilized SCoT markers to evaluate the genetic diversity and relationships among Aegilops and Triticum species. The study revealed high genetic variation within species, particularly in Ae. cylindrica, and demonstrated the effectiveness of SCoT markers in grouping samples based on their genomic constitutions. This analysis supports the hypothesis that T. urartu and Ae. tauschii are diploid ancestors of T. aestivum, providing valuable insights for wheat breeding and genetic improvement (Ghobadi et al., 2021; Pour-Aboughadareh et al., 2018).

Ghobadi et al. (2021) used SCoT and CBDP markers to estimate genetic diversity and population structure in Triticum aestivum and Aegilops species. The study found that both marker systems were effective in evaluating genetic diversity, with SCoT markers showing higher polymorphism. The results indicated significant genetic variation within species, with Ae. cylindrica exhibiting the highest genetic diversity. These findings highlight the potential of SCoT and CBDP markers in genetic studies and breeding programs for wheat and its wild relatives (Pour-Aboughadareh et al., 2021).

5.3 SCoT and CBDP markers used to estimate genetic diversity and population structure

In a comprehensive study conducted by Ghobadi et al. (2021), SCoT and CBDP markers were used to evaluate the genetic diversity (Figure 2) and population structure of wheat and its wild relatives. The study involved 91 samples, covering species including common wheat (Triticum aestivum), jointed goatgrass (Aegilops cylindrica), and goatgrass (Aegilops crassa). Figure 2 shows that the within-species variation accounted for 78% and the between-species variation accounted for 22% with SCoT markers; for CBDP markers, within-species variation accounted for 80% and between-species variation accounted for 20%. This indicates that SCoT markers exhibit higher interspecies resolution in detecting genetic diversity, highlighting their effectiveness and advantage in genetic diversity studies within the Triticeae crops.

Both the SCoT and CBDP marker systems generated a large number of polymorphic fragments, indicating significant genetic variation. Analysis of molecular variance (AMOVA) revealed that genetic diversity within species was higher than that between species. Therefore, SCoT and CBDP markers are effective tools for assessing genetic diversity and contributing to the genetic improvement of wheat and its wild relatives (Pour-Aboughadareh et al., 2021).

6 Challenges and Limitations

6.1 Technical challenges in using SEM for genetic studies

Scanning Electron Microscopy (SEM) has been instrumental in providing high-resolution images for the examination of grain surface sculptures in Triticeae species. However, several technical challenges persist. One significant challenge is the preparation of samples, which requires meticulous handling to avoid artifacts that could mislead the interpretation of surface structures. Additionally, SEM analysis is time-consuming and requires specialized equipment and expertise, which may not be readily available in all research settings. The resolution and depth of field, while generally high, can sometimes be insufficient for distinguishing very fine genetic differences, limiting its utility in comprehensive genetic studies (Mohamed et al., 2017).

6.2 Limitations of SCoT markers

Start Codon Targeted (SCoT) markers, while useful for genetic diversity studies, have their limitations. One major limitation is their dominant nature, which means they cannot distinguish between homozygous and heterozygous loci, potentially leading to an underestimation of genetic diversity. Additionally, the reproducibility of SCoT markers can be inconsistent, influenced by factors such as primer length and annealing temperature, which complicates their use in large-scale studies (Collard and Mackill, 2009). The polymorphic information content (PIC) and resolving power (Rp) of SCoT markers, although generally adequate, are often lower compared to other marker systems like CAAT-box derived polymorphism (CBDP) markers, which limits their effectiveness in certain applications (Nosair, 2020; Ghobadi et al., 2021).

6.3 Addressing the challenges: potential solutions and future directions

To address the technical challenges associated with SEM, advancements in sample preparation techniques and the development of more user-friendly SEM equipment could be beneficial. Training programs to enhance the expertise of researchers in SEM technology would also help mitigate some of the current limitations (Mohamed et al., 2017).

For SCoT markers, combining them with other marker systems such as CBDP and SSR markers can enhance the robustness and reliability of genetic studies. This combined approach leverages the strengths of each marker system, providing a more comprehensive analysis of genetic diversity and population structure (Figure 3) (Ghobadi et al., 2021; Pour-Aboughadareh et al., 2022). In a study by Pour-Aboughadareh et al. (2022), a dendrogram was generated using four marker systems (SCoT, CBDP, SSR, and combined data) to reveal the phylogenetic relationships between the studied bread wheat and its wild relatives. Comparing the dendrograms generated by these marker systems, it was observed that SCoT markers, when combined with other marker systems, demonstrated a high degree of consistency and stability. Particularly in population classification and genetic diversity analysis, SCoT markers provided reliable resolution, and when combined with other markers, they allowed for a more comprehensive and accurate understanding of the genetic background of Triticeae crops. This multi-marker combination enhanced the robustness of the study, leading to a deeper understanding of the genetic relationships among different varieties and species.

Future research should focus on optimizing the conditions for SCoT marker amplification to improve its reproducibility and explore the development of co-dominant markers that can provide more detailed genetic information (Collard and Mackill, 2009; Nosair, 2020).

7 Future Prospects and Technological Advancements

7.1 Emerging technologies in SEM and their potential applications

Scanning Electron Microscopy (SEM) has been instrumental in providing high-resolution imaging for the examination of grain surface sculptures in Triticeae species. Recent advancements in SEM technology, such as enhanced imaging resolution and the ability to perform three-dimensional reconstructions, have significantly improved the accuracy and detail of morphological studies. These advancements allow for more precise discrimination and identification of cultivars, as demonstrated in the study of Northern African Triticum aestivum L. cultivars, where SEM revealed distinct grain surface sculptures that were crucial for taxonomic and genetic characterization (Mohamed et al., 2017). Future applications of SEM could include automated image analysis and machine learning algorithms to further enhance the identification and classification processes.

7.2 Advances in SCoT marker development

Start Codon Targeted (SCoT) markers have emerged as a powerful tool for genetic analysis due to their ability to generate gene-targeted polymorphisms. Recent studies have shown that SCoT markers are highly effective in assessing genetic diversity and population structure in various Triticeae species. For instance, SCoT markers were used to analyze genetic variability in wheat germplasm, revealing significant polymorphism and high genetic diversity within species (Ghobadi et al., 2021; Pour-Aboughadareh et al., 2022). Additionally, the development of SCoT markers has facilitated the identification of specific genetic traits, such as those related to environmental stress tolerance and agronomic performance (Etminan et al., 2016; Pour-Aboughadareh et al., 2018). The continuous improvement of SCoT marker techniques, including the development of co-dominant markers and their integration with other molecular markers, promises to enhance the precision and efficiency of genetic studies in Triticeae.

7.3 Integration of advanced tools for enhanced genetic improvement

The integration of advanced molecular tools, such as SCoT markers, with traditional breeding methods and other molecular markers like SSRs and CBDPs, offers a comprehensive approach to genetic improvement in Triticeae. Studies have shown that combining different marker systems can provide a more detailed understanding of genetic relationships and diversity. For example, the use of SCoT and CBDP markers together has been shown to be effective in genetic fingerprinting and fine mapping in wheat and its wild relatives (Gholamian et al., 2019; Pour-Aboughadareh et al., 2022). Moreover, the development of sequence-tagged microsatellites (STMs) has increased marker density on genetic maps, further enhancing marker-assisted selection and breeding programs (Hayden et al., 2006). The future of genetic improvement in Triticeae lies in the seamless integration of these advanced tools, enabling more efficient and targeted breeding strategies to develop cultivars with improved traits and resilience to environmental stresses.

8 Concluding Remarks

The utilization of Start Codon Targeted (SCoT) markers and Structural Equation Modeling (SEM) has shown significant promise in the genetic improvement of Triticeae. SCoT markers have been effectively used to assess genetic diversity, population structure, and genetic relationships in various plant species, including Triticeae. For instance, SCoT markers have demonstrated high polymorphism and reproducibility, making them suitable for genetic analysis and breeding programs. Studies have shown that SCoT markers can differentiate between toxic and non-toxic accessions, as well as between wild and domesticated populations, highlighting their utility in identifying genetic variations and potential for crop improvement. Additionally, the combination of SCoT and other marker systems, such as ISSR and CBDP, has provided comprehensive insights into genetic diversity and population structure, further supporting their application in genetic research.

The future of genetic research in Triticeae looks promising with the continued application and development of SCoT markers. These markers offer a cost-effective, reliable, and efficient method for genetic analysis, which is crucial for the improvement of Triticeae crops. The ability of SCoT markers to generate gene-targeted polymorphisms and their high reproducibility make them invaluable tools for genetic mapping, marker-assisted selection, and the identification of desirable traits. Furthermore, the integration of SCoT markers with advanced genomic technologies and bioinformatics tools will enhance our understanding of the genetic basis of important agronomic traits, leading to the development of superior Triticeae varieties.

To fully realize the potential of SCoT markers in Triticeae genetic improvement, continued research and innovation are essential. Researchers should focus on expanding the application of SCoT markers to a broader range of Triticeae species and exploring their potential in combination with other molecular markers and genomic tools. Collaborative efforts between geneticists, breeders, and bioinformaticians will be crucial in developing comprehensive genetic databases and breeding programs that leverage the power of SCoT markers. Additionally, investment in training and capacity building for researchers in developing regions will ensure the widespread adoption and effective use of these markers in Triticeae improvement programs. By fostering a collaborative and innovative research environment, we can accelerate the genetic improvement of Triticeae, ensuring food security and agricultural sustainability for future generations.

Acknowledgments

Thank you to the peer reviewers for their suggestions on improving this study.

Conflict of Interest Disclosure

The author affirms that this research was conducted without any commercial or financial relationships that could be construed as a potential conflict of interest.

References

Amom T., Tikendra L., Apana N., Goutam M., Sonia P., Koijam A., Potshangbam A., Rahaman H., and Nongdam P., 2020, Efficiency of RAPD, ISSR, iPBS, SCoT and phytochemical markers in the genetic relationship study of five native and economical important bamboos of North-East India, Phytochemistry, 174: 112330.

https://doi.org/10.1016/j.phytochem.2020.112330

PMid:32146386

Bauer E., Schmutzer T., Barilar I., Mascher M., Gundlach H., Martis M., Twardziok S., Hackauf B., Gordillo A., Wilde P., Schmidt M., Korzun V., Mayer K., Schmid K., Schön C., and Scholz U., 2017, Towards a whole‐genome sequence for rye (Secale cereale L.), The Plant Journal, 89: 853-869.

https://doi.org/10.1111/tpj.13436

PMid:27888547

Bobik K., Dunlap J., and Burch-Smith T., 2014, Tandem high-pressure freezing and quick freeze substitution of plant tissues for transmission electron microscopy, Journal of visualized experiments: JoVE, 92: e51844.

https://doi.org/10.3791/51844

PMid:25350384 PMCid:PMC4692431

Collard B., and Mackill D., 2008, Marker-assisted selection: an approach for precision plant breeding in the twenty-first century, Philosophical Transactions of the Royal Society B: Biological Sciences, 363: 557-572.

https://doi.org/10.1098/rstb.2007.2170

PMid:17715053 PMCid:PMC2610170

Collard B., and Mackill D., 2009, Start codon targeted (SCoT) polymorphism: a simple, novel DNA marker technique for generating gene-targeted markers in plants, Plant Molecular Biology Reporter, 27: 86-93.

https://doi.org/10.1007/s11105-008-0060-5

Elshire R., Glaubitz J., Sun Q., Poland J., Kawamoto K., Buckler E., and Mitchell S., 2011, A robust, simple genotyping-by-sequencing (GBS) approach for high diversity species, PLoS ONE, 6(5): e19379.

https://doi.org/10.1371/journal.pone.0019379

PMid:21573248 PMCid:PMC3087801

Etminan A., Pour-Aboughadareh A., Mohammadi R., Ahmadi-Rad A., Noori A., Mahdavian Z., and Moradi Z., 2016, Applicability of start codon targeted (SCoT) and inter-simple sequence repeat (ISSR) markers for genetic diversity analysis in durum wheat genotypes, Biotechnology & Biotechnological Equipment, 30: 1075-1081.

https://doi.org/10.1080/13102818.2016.1228478

Feng S., He R., Yang S., Chen Z., Jiang M., Lu J., and Wang H., 2015, Start codon targeted (SCoT) and target region amplification polymorphism (TRAP) for evaluating the genetic relationship of Dendrobium species, Gene, 567(2): 182-188.

https://doi.org/10.1016/j.gene.2015.04.076

PMid:25936992

Ghobadi G., Etminan A., Mehrabi A., and Shooshtari L., 2021, Molecular diversity analysis in hexaploid wheat (Triticum aestivum L.) and two Aegilops species (Aegilops crassa and Aegilops cylindrica) using CBDP and SCoT markers, Journal of Genetic Engineering & Biotechnology, 19(1): 56.

https://doi.org/10.1186/s43141-021-00157-8

PMid:33852105 PMCid:PMC8046865

Gholamian F., Etminan A., Changizi M., Khaghani S., and Gomarian M., 2019, Assessment of genetic diversity in Triticum urartu Thumanjan ex Gandilyan accessions using start codon targeted polymorphism (SCoT) and CAAT-box derived polymorphism (CBDP) markers, Biotechnology & Biotechnological Equipment, 33: 1653-1662.

https://doi.org/10.1080/13102818.2019.1691466

Habiba R., Bashasha J., Haffez S., and Leilah A., 2021, Assessment of genetic diversity using SCoT markers and some morphological traits in ten lines of barley (Hordeum vulgare L.), Assiut Journal of Agricultural Sciences, 52(4): 53-65.

https://doi.org/10.21608/ajas.2022.111745.1077

Hayden M., Stephenson P., Logojan A., Khatkar D., Rogers C., Elsden J., Koebner R., Snape J., and Sharp P., 2006, Development and genetic mapping of sequence-tagged microsatellites (STMs) in bread wheat (Triticum aestivum L.), Theoretical and Applied Genetics, 113: 1271-1281.

https://doi.org/10.1007/s00122-006-0381-4

PMid:16932882

Hyun D., Sebastin R., Lee K., Lee G., Shin M., Kim S., Lee J., and Cho G., 2020, Genotyping-by-sequencing derived single nucleotide polymorphisms provide the first well-resolved phylogeny for the genus Triticum (Poaceae), Frontiers in Plant Science, 11: 688.

https://doi.org/10.3389/fpls.2020.00688

PMid:32625218 PMCid:PMC7311657

Ibrahim M., Mohamed A., Teleb S., Ibrahim S., and Tantawy M., 2017, Taxonomic and molecular study on some Asian cultivars of Triticum aestivum L., Taeckholmia, 37(1): 16-29.

https://doi.org/10.21608/taec.2017.11932

Igwe D., Ihearahu O., Osano A., Acquaah G., and Ude G., 2021, Assessment of genetic diversity of Musa species accessions with variable genomes using ISSR and SCoT markers, Genetic Resources and Crop Evolution, 69: 49-70.

https://doi.org/10.1007/s10722-021-01202-8

Kuipers J., Boer P., and Giepmans B., 2015, Scanning EM of non-heavy metal stained biosamples: large-field of view, high contrast and highly efficient immunolabeling, Experimental Cell Research, 337(2): 202-207.

https://doi.org/10.1016/j.yexcr.2015.07.012

PMid:26272543

Lenthe W., Stinville J., Echlin M., Chen Z., Daly S., and Pollock T., 2018, Advanced detector signal acquisition and electron beam scanning for high resolution SEM imaging, Ultramicroscopy, 195: 93-100.

https://doi.org/10.1016/j.ultramic.2018.08.025

PMid:30216796

Lu B., and Ellstrand N., 2014, World food security and the tribe Triticeae (Poaceae): genetic resources of cultivated, wild, and weedy taxa for crop improvement, Journal of Systematics and Evolution, 52(6): 661-666.

https://doi.org/10.1111/jse.12131

Mascher M., Gundlach H., Himmelbach A., Beier S., Twardziok S., Wicker T., Radchuk V., Dockter C., Hedley P., Russell J., Bayer M., Ramsay L., Liu H., Haberer G., Zhang X., Zhang Q., Barrero R., Li L., Taudien S., Groth M., Felder M., Hastie A., Šimková H., Staňková H., Vrána J., Chan S., Muñoz‐Amatriaín M., Ounit R., Wanamaker S., Bolser D., Colmsee C., Schmutzer T., Aliyeva-Schnorr L., Grasso S., Tanskanen J., Chailyan A., Sampath D., Heavens D., Clissold L., Cao S., Chapman B., Dai F., Han Y., Li H., Li X., Lin C., McCooke J., Tan C., Wang P., Wang S., Yin S., Zhou G., Poland J., Bellgard M., Borisjuk L., Houben A., Doležel J., Ayling S., Lonardi S., Kersey P., Langridge P., Muehlbauer G., Clark M., Cáccamo M., Schulman A., Mayer K., Platzer M., Close T., Scholz U., Hansson M., Zhang G., Braumann I., Spannagl M., Li C., Waugh R., and Stein N., 2017, A chromosome conformation capture ordered sequence of the barley genome, Nature, 544: 427-433.

https://doi.org/10.1038/nature22043

PMid:28447635

Mochida K., Saisho D., Yoshida T., Sakurai T., and Shinozaki K., 2008, TriMEDB: A database to integrate transcribed markers and facilitate genetic studies of the tribe Triticeae, BMC Plant Biology, 8: 72.

https://doi.org/10.1186/1471-2229-8-72

PMid:18590523 PMCid:PMC2474609

Mohamed A., Ibrahim M., Teleb S., and Tantawy M., 2017, SEM and SCoT Markers Unveil New taxonomic and genetic insights about some Northern African Triticum aestivum L. cultivars, Vegetos, 30: 34.

https://doi.org/10.5958/2229-4473.2017.00006.4

Nadeem M., Nawaz M., Shahid M., Doğan Y., Comertpay G., Yıldız M., Hatipoğlu R., Ahmad F., Alsaleh A., Labhane N., Özkan H., Chung G., and Baloch F., 2018, DNA molecular markers in plant breeding: current status and recent advancements in genomic selection and genome editing, Biotechnology & Biotechnological Equipment, 32: 261-285.

https://doi.org/10.1080/13102818.2017.1400401

Nosair H., 2020, Genetic diversity studies on seven Egyptian wheat (Triticum aestivum) cultivars using Scot and ISSR polymorphism markers, Taeckholmia, 40(1): 143-151.

https://doi.org/10.21608/taec.2020.39905.1025

Pease R., 2008, Significant Advances in Scanning Electron Microscopes (1965-2007), Advances in Imaging and Electron Physics, 150: 53-86.

https://doi.org/10.1016/S1076-5670(07)00002-X

Pour-Aboughadareh A., Ahmadi J., Mehrabi A., Etminan A., and Moghaddam M., 2018, Insight into the genetic variability analysis and relationships among some Aegilops and Triticum species, as genome progenitors of bread wheat, using SCoT markers, Plant Biosystems - An International Journal Dealing with all Aspects of Plant Biology, 152: 694-703.

https://doi.org/10.1080/11263504.2017.1320311

Pour-Aboughadareh A., Poczai P., Etminan A., Jadidi O., Kianersi F., and Shooshtari L., 2022, An analysis of genetic variability and population structure in wheat germplasm using microsatellite and gene-based markers, Plants, 11(9): 1205

https://doi.org/10.3390/plants11091205

PMid:35567205 PMCid:PMC9103345

Schroeder-Reiter E., Pérez-Willard F., Zeile U., and Wanner G., 2009, Focused ion beam (FIB) combined with high resolution scanning electron microscopy: a promising tool for 3D analysis of chromosome architecture, Journal of Structural Biology, 165(2): 97-106.

https://doi.org/10.1016/j.jsb.2008.10.002

PMid:19059341

Spehner D., Steyer A., Bertinetti L., Orlov I., Benoit L., Pernet-Gallay K., Schertel A., and Schultz P., 2020, Cryo-FIB-SEM as a promising tool for localizing proteins in 3D, Journal of Structural Biology, 211(1): 107528.

https://doi.org/10.1016/j.jsb.2020.107528

PMid:32387573

Souza W., and Attias M., 2018, New advances in scanning microscopy and its application to study parasitic protozoa, Experimental Parasitology, 190: 10-33.

https://doi.org/10.1016/j.exppara.2018.04.018

PMid:29702111

Uryu K., Soplop N., Sheahan T., Catanese M., Huynh C., Pena J., Boudreau N., Matei I., Kenific C., Hashimoto A., Hoshino A., Rice C., and Lyden D., 2023, Advancement in cellular topographic and nanoparticle capture imaging by high resolution microscopy incorporating a freeze-drying and gaseous nitrogen-based approach, bioRxiv, 2023-12.

https://doi.org/10.1101/2023.09.28.559906