Detected Genetic Markers for Three Varieties of Rice ( Oryza sativa L.) under Nano-Particles

Despite the enormous advancements in breeding, fresh and accessible methods of plant development are preferred. Many research have been done to determine the impact of zinc oxide nanoparticles (ZnO NPs) and magnesium nanoparticles (MgONPs) on plant growth. ZnO NPs and MgONPs have been used extensively in agriculture. Few research have looked into the potential impact of ZnO NPs on grain Zn content or cereal yield formation to date. The goal of the study was to demonstrate how varied concentrations of nanoparticles affected root germination in three different rice types (Giza 177, Sakha super 300, and Sakha 108) and the fungus Fusariummoniliforme, which causes foot rot in rice. Here, we performed a pot experiment


Introduction
For the majority of the world, rice (Oryza sativa L.) is an essential cereal food crop (Khush, 2005;Khan et al., 2022).Being heavy in carbs, low in fat, and abundant in calories, protein, and vitamins gives it great nutritional advantages (El-Mowafi et al., 2021).Its area is almost 162 million hectares, and it will produce 756 million tonnes by the year 2022 (Faostat).Additionally, in order to deal with the threat of environmental challenges and the ongoing population growth, this production should be enhanced (Fageria, 2007).
The development of nanotechnology has provided favourable applications for sustainable agriculture.The agriculture sector faces numerous and unprecedented challenges, such as decreased crop yields due to biotic and abiotic pressures, including nutrient deficiencies and pathogenic bacteria.It is a relatively new practise to utilise nanoparticles to enhance plant development and reduce plant diseases (Rastogi et al., 2019).However, the problem of phytotoxicity brought on by NP exposure remains unclear.Recent studies (Youssef and Elamawi 2018) revealed a considerable effort to understand how nanoparticles affect plant growth and suggested a favourable impact of NPs on plant development with the potential to be employed as nano-fertilizer in the future.More than half of the world's population depends heavily on rice (Oryza sativa L.), therefore to keep up with expanding demand, rice production needs to double by 2050 (FAO 2009).
Because of the seed's abundance in proteins, minerals, vitamins, and fibres, it is also regarded as the primary diet for all human ages.Magnaporthe oryzae Couch, also known as Pyricularia oryzae Cavara, is the organism that causes rice blast (Couch and Kohn 2002).Over 50 kinds of grass are attacked by M. oryzae, which is present in about 80 countries (Ou 1985).According to Nalley et al. (2016), it causes around 10-30% of the world's rice crop to be lost.The majority of rice cultivars are vulnerable to several fungal races.Fungicides are frequently used to control blast, but they are losing favour because they raise the risk of M. oryzae developing fungicide resistance and because they run afoul of public concerns about the effects of fungicide residues on human health and the environment.
Controlling the disease is difficult because the fungus is so varied (Ou 1985).Using nanoparticles as an alternative could help regulate this illness.According to Young et  When used as a fertiliser, ZnO NPs may be more readily available to plants than standard Zn fertiliser, which could result in improved rice growth.According to Kulhare et al. (2017), foliar Zn fertiliser application increased rice productivity and grain Zn concentration.Due to its special qualities, such as biodegradability, non-toxicity, suppression of biofilm growth, degradation of hazardous colours such as methyl violet, and many more (Fernandes, et al., 2020), magnesium oxide nanoparticles have become a promising contender for solving a variety of problems.
In the current work, ISSR markers have been employed to ascertain the genetic diversity among various Hassawi rice landraces.In order to produce polymorphic DNA bands in the rice genome, the effectiveness of ISSR primers of di-and tri-nucleotide repeats has been investigated (Al-Turki et al., 2015).
The outcomes demonstrated that RAPD primers generated 49 bands with a size range of 0.1-3 kb and an 87.75% polymorphism proportion.For RAPD, 43 polymorphic bands with distinguishable bands were discovered.According to morphological traits and RAPD analysis, the UPGMA Dendrogram was classified into three groups (Rashad et al., 2023).determining how various medium compositions affect the regrowth and embryogenic responses of various barley genotypes.The three barley genotypes El-kasr, G126, and G130 were successfully evaluated for the presence of somaclonal variation using biochemical and molecular genetics examinations of protein, isozymes, and RAPD-PCR (Rashad et al., 2020;Rashad et al., 2023).the capacity of RAPD markers to identify relationships between diverse genotypes and demonstrate how yield metrics at the molecular level are related.In order for barley breeders working with gene banks to ensure that it can be produced sustainably and utilised to its fullest extent, it is imperative to understand the diversity of barley.Using cutting-edge molecular, biochemical, and physiological techniques, this can be accomplished by thoroughly phenotyping and genotyping the barley collections (Merwad et al., 2020).ISSR analysis and physical features can be used to find genetic variations (Shata et al., 2021).SSR marker analysis is less effective than ISSR markers at detecting genetic diversity among the evaluated cultivars of Alfalfa.In comparison to the SSR marker, the ISSR marker is more precise and yields more data (Heiba et al., 2022).ISSR primers produced 37 bands with widths ranging from 100 to 2000 bp and an 87.5% success rate, proportion of polymorphism.Polymorphic information content (PIC) for ISSR was 0.74.The UPGMA Dendrogram was divided into two clusters based on physical characteristics, and ISSR analysis is a helpful tool for locating genetic variations, Rashad et al 2022.
Using molecular markers ISSR and RAPD, the current study aims to assess the effects of ZnO and MgO nanoparticles on the germination of three different rice varieties (Giza 177, Sakha super 300, and Sakha 108) and the fungal activity Fusariummoniliforme in rice that causes foot rot.Additionally, to research the potential impacts of varying ZnO NP application rates as a nano-fertilizer and anti-fungal agent to reduce rice Fusarium moniliforme and increase cultivar grain productions.

Materials And Methods
Synthesis of Zinc and Magnesium Nanoparticles from different precursor 90 ml of 10 mM zinc sulphate, magnesium sulphate, and magnesium chloride were added gradually to 10 ml of 25% Eucalyptus melliodora (camphor) aqueous extract in a 250 ml Erlenmeyer flask.For 30 minutes, the mixture was heated on a magnetic stirrer at 80 °C while being continuously stirred.After a while, NPs started to form and the colour changed from white to greyish white.For 30 minutes, 6000 rpm of centrifugation was used to separate the nanoparticles.After discarding the supernatant, the pellet underwent three centrifugal washes in distilled water.After being dried for 24 hours at 60 °C in a vacuum oven, the residue powder was characterised using a variety of methods.

XRD and UV-Vis Spectra Analysis
Spectra were analysed using a double-beam spectrophotometer (model 8453E, HP).After dilution into deionized water, a tiny aliquot of the NPs solution was used to measure the UV-Vis spectrum of the reaction solution in the 200-600 nm wavelength range.At room temperature, the solution was examined after being pipetted into a test tube.

Transmission Electron Microscope (TEM)
A transmission electron microscope (JEOL 100CX II, Japan, with an accelerating voltage of 100 kV) was used to examine several NPs.Drops of an aqueous solution of NPs were placed on carbon-coated copper grids to create nanoparticles for TEM investigation, which were then allowed to dry for five minutes before the excess solution was blotted away with blotting paper at room temperature.

* Fourier Transform Infrared Spectroscopy (FT-IR)
FT-IR spectra were analysed using a data station-equipped BOMEM FT-IR Spectrometer (MB 147, Canada) in the range of 4000 to 400 cm-1.About 100 mg of dried samples and 100 mg of spectral grade KBr were combined, and then the mixture was pressed into discs using hydraulic pressure.

In vitro fungal activity and seed germination test
• Three different rice types, Giza 177, Sakha super300, and Sakha 108, were employed in the experiment.In each of the three huge petri dishes, 75 seeds of each variety were planted, with 25 seeds for each Nano-treatment.The first treatment used a Nano-Zink solution, which is represented by the letter (A).The second treatment used a high concentration Nano-magnesium solution, which is represented by the letter (B).The third treatment used a low concentration of Nano-magnesium solution, which is represented by the letter (D).
• Distilled water was used as the first irrigation for all experimental varieties.The treatments under study were then irrigated for a period of fifteen days using a regulator watering system, and the following measurements were made: the length of the shoot, the length of the roots, and the percentage of germination.
To investigate the impact of nanoparticles at various concentrations on rice plant seed germination.seeds that were exposed to solutions containing Zn and Mg nanoparticles at concentrations of 10, 20, and 100 ppm.To examine the effectiveness of synthesised nanoparticles as an antifungal agent, PDA Available online at: https://jazindia.com-938 -plates were also treated with varied doses of 10, 20, and 100 ppm of Zn and Mg nanoparticle solution before being inoculated with Fusariummoniliforme

The isolation and cultivation of the fungus:
Rice seeds were sterilised for two minutes by being submerged in a solution containing 0.5% sodium hypochlorite (NaOH) in order to cultivate and isolate the fungus.A sample is placed on a medium made of potato dextrose agar (PDA) after being washed twice with sterile distilled water.The plates underwent a 5-to 7-day incubation period at a temperature of 25°C.The growing fungus were then refined via the single-spore method.

Identification:
Using the key of imperfect fungi (Barnett and Hunter, 1972;Nelson et al., 1983), the isolates of the fungus were identified based on their morphological traits and microscopic examination.The fungal isolate was placed into glass flasks containing 100 mL of potato dextrose broth prior to DNA extraction.The flasks were then continuously shaken until mycelium had developed.To get rid of any traces of the culture medium, the mycelia were collected, filtered through cheesecloth, and then repeatedly washed with sterile distilled water.
Using a Precellys homogenizer, 50 mg of fresh mycelia from the fungal isolates were mechanically broken apart in order to extract the DNA and do PCR sequencing.The Jena Bioscience DNA extraction kit was then used to extract the entire sample's DNA.Following a dilution procedure to isolate the DNA from its surrounds, the nuclear ribosomal DNA's ITS sections were amplified using the primers ITS1F-4, as explained by Gardes and Bruns in 1993.The ultimate volume of the polymerase chain reaction (PCR) was 20 L. The reaction mixture included 10 L of the amaR One PCRTM Gene DireX, Inc., 1 M of each primer, and 50 ng of template DNA.Mix master.The thermal cycling settings included an initial denaturation phase lasting two minutes at 94 °C, followed by 35 cycles of denaturation lasting one minute at 94 °C, annealing for 30 seconds at 58 °C, and extension lasting one minute at 72 °C.A final extension step was carried out for seven minutes at 72 °C.The results of the PCR were examined under UV light after electrophoresis on a 2% agarose gel coloured with Gel-Red (Biotium TM).With the aid of a solution containing 1 volume of 20% polyethylene glycol 8000 and 2.5 M NaCl, the PCR products were purified by precipitation.The primers ITS1F(5′-CTT GGT CAT TTA GAG GAA GTAA-3′) and ITS4(5′-TCC TCC GCT TGA TAT GC-3′) were used to perform forward and reverse sequencing on the PCR products that had undergone purification, in accordance with White et al. (1990).

DNA extraction of ISSR and RAPD reaction conditions:
DNA extraction: using DNA kit methods PCR results were shot using the Biometra -Bio Documentations gel documentation system and UV transilluminated using a 100 bp DNA ladder.According to the gel analyzer technique, the amplified bands were graded as (1) for all studied wheat types' presence and (0) for their absence.A list of 10 primers is shown in Table 1 for the purpose of finding polymorphism among the three rice cultivars.These primers were developed by ISSR-PCR in accordance with Williams et al.'s (1990) guidelines with a few minor modifications.
In PCR-ISSR tests, inter-simple sequence repeats (ISSRs) are utilized to identify ISSRs.The reaction mixture, which consists of 20 ml of standardization (20 ml of PCR buffer 1X, 2.5 mM MgCl2, 1 mM dNTPs, 10 pmol primer, 1-unit Taq polymerase, and 50 ng genomic DNA; 38 cycles of 56 °C: 1 min annealing, 2 min extension, and 10 min final extension at 72 °C).The ISSR-PCR data will be analysed using a 1.4% (w/v) agarose gel.Heiba et al. (2022) will assess the size of the amplified bands using a 1Kb and 100bpDNA ladder from Ferments Life Sciences and a gel documentation device (SyngeneTM) to capture pictures of the gels.

ISSR-PCR analysis:
In PCR tests, ISSR primers will be utilized, according to Zietkiewciz et al. (1994).The extracted DNA will be used to run an ISSR-PCR reaction on numerous samples.The reaction mixture will consist of 2 l of genomic DNA, 1 l of the primer, 2.5 l of 10X Taq DNA polymerase reaction buffer, 1.5 units of Taq DNA polymerase, and 200 mM of each dNTP.The amplifications will be performed using a DNA amplification Thermo cycler (PTC-100 PCR version 9.0 from M J Research-USA).The equipment is set up to work in the following circumstances: a 5 min denaturation step at 94°C, followed by 35 cycles of 30 s at 94°C, 90 s at the annealing temperature (specified by each primer), and 90 s at 72°C.For the amplifications, which will be performed at least twice in order to undertake further data analysis, only repeatable products will be used.
Electrophoresis on a gel using a DNA ladder (1Kb) in 1X TAE buffer, the ISSR amplification products will be separated on 1.5% agarose gels.The gels will next be identified by staining with ethidium bromide, in accordance with Sambrook et al., (1989).The PCR findings will be imaged and documented using the Biometra -Bio Doc.Analyze gel documentation system.

Electrophoresis and RAPD Analysis
Using five enhanced RAPD primers, the DNA of the sunflower germplasm was amplified using a polymerase chain reaction (PCR).To determine the genetic diversity among the sunflower varieties, RAPDs were utilized.Denaturation at 95°C for one minute, annealing at 36°C for one minute, and extension at 72°C for two minutes for 35 cycles comprised the three phases of the PCR.The PCR programmed was set up to keep the outcomes at 4°C. 6 l of loading dye (0.50% xylene cyanol and 0.50% bromophenol blue) were added to the goods and spun in a mini centrifuge.The PCR result was electrophoresed on a 2% agarose gel at a voltage of 100 V, and amplified bands were found on a gel documentation system by Bio-Rad (Hercules) (Rashad et al., 2020).
Data analysis: Only clear, unambiguous, and reproducible bands will be considered for data analysis.Every band will be handled individually as a locus.
Data will be rated as (1) for presence and (0) for absence for each of the seven samples of barley using the Gel analyser programme, which is used to identify positive and negative markers.In order to make a dendrogram using the unweight pair group approach with arithmetical average (UPGMA), the similarity coefficients will be generated by the SPSS programme version 10 (Norman et al., 1975).

Statistical Analysis
The data were statistically analysed in accordance with Gomez and Gomez (1984).The least significant differences (LSD) were used to compare treatment mean differences at the 5% and 1% levels of probability.

Characterization of Zn and Mg nanoparticles.
Zn particle synthesis was validated by UV-Vis spectra analysis and X-ray diffraction (XRD), and UV-Vis spectra analysis was carried out using a double-beam spectrophotometer (8453E, HP).By monitoring the UV-Vis spectra of the reaction solution in the 200-600 nm wavelength range and the ZnNPs absorption peak at 310 nm wavelength (Fig. 1), the solution of NPs.nanoparticles was seen.While the peak at 1613cm-1 is attributed to C=O stretching in amides (tertiary amides) or N-H bend from amides (primary amide), possibly similar to amino acids or protein, the peak at 1063cm-1, from 605 to 429, suggests the ZnO.Potential secondary metabolites include tannins, saponins, terpinoids, and terpenes.The distinct peaks at 2103, 1437, 984, and 441 with precursor MgOac and 984cm-1 with MgOsu suggest the MgO, in addition to the emergence of the MgONPs synthesised from sulphate or acetate (MgOsu and MgOac) displayed in Fig. 3B.

In vitro seed germination
The effects of nanoparticles at various concentrations on rice plant root germination are depicted in this figure.seeds treated with solutions containing Zn and Mg nanoparticles in concentrations of 10, 20, and 100 ppm.
Table 1 shows a significant increase in length when compared to control, but Figure 3 shows a significant decrease in plant length when Zn and Mg-NP concentration was raised.This indicates that while Zn and Mg nanoparticles are beneficial to plants at certain concentrations, raising the concentration has toxic effects on plant growth.

The isolation identification and cultivation of the fungus:
Fusariummoniliforme was identified and confirmed by DNA sequenced and submitted by accession number OR523600.
When Zn and Mg nanoparticle concentration was increased, however, plant length significantly decreased, as shown in Fig. 4.This shows that while Zn and Mg nanoparticles have positive effects on the plant at certain concentrations, when that concentration is increased, this leads to toxic effects on plant growth, as shown in Fig. 3. Fungal growth has significantly decreased when Zn and Mg-NPs(ac) concentration was increased, as shown in Fig. 1.This indicates that Zn, and Mg nanoparticles have positive effects on the fungal growth when at all concentrations; however, when that concentration is increased, this has an unfavorable impact on the environment.
According to Petosa et al. (2017), nanoparticles of various metal oxides can play a significant role in promoting plant development and yield as well as suppressing plant diseases.In the current study, it was found that, depending on the concentration and timing of applications, Zn NPs are more frequently linked to improvements in rice growth and yield when cultivated in pathogen-infected soil.All of the findings were unexpected considering that immature seedlings at the nursery were given a single dose Detected Genetic Markers for Three Varieties of Rice (Oryza sativa L.) under Nano-Particles Available online at: https://jazindia.com-942 -to have a long-lasting, frequently season-long effect.These results are crucial for future field applications because they allow for the safe and regulated delivery of modest amounts of NPs to early seedlings, all while easing worries about NP exposure to both humans and the environment.

Effect of ZnO NPs on genomic DNA of Rice (Oryza sativa L.) using ISSR and RAPD-PCR primers:
In order to ascertain the impact of the various doses of ZnO-NPs and MgONPs utilised in this investigation, the genomic DNA of rice (Oryza sativa L.) was subjected to seven ISSR markers and three RAPD primers (Table 1 and Figures 5&6&7).Each and every one of the ISSR primers amplified and produced polymorphic alleles that displayed monomorphic alleles.In the meantime, three of the RAPD-PCR primers employed in this investigation amplified and produced polymorphic bands.

ISSR markers:
Five bands altogether, ranging in size from 383 to 1084 bp, were produced as a result of the samples' DNA being amplified using UBC809.There was only one polymorphism with distinct bands found.The amount of polymorphism was 20%; four monomorphic bands and a single band, identified at a molecular size of 477 bp for the Giza 177, occurred in the sample treated with 10, 20, and 100 mg/L ZnO-NPs and MgONPs, respectively.In the instance of the UBC823 primer, there were a total of eight bands; all of them displayed four monomorphic bands and ranged in size from 349 to 1403 bp.There were also two distinct bands, which were found at molecular sizes of 349 bp and 928 bp for the Giza 177.Therefore, 50% of the population was polymorphic.Ten bands in all, spanning 303 to 1377 bp, were detected for the UBC807 primer (four polymorphic, six monomorphic, and 40% polymorphic).
While being missing at all concentrations of ZnO NPs and MgONPs, two distinct bands with molecular sizes of 341 bp for the Sakha super 300 and 1130 bp for the Sakha 108 were found.A 42.86% polymorphism rate was present.Seven bands in all, ranging in size from 478 to 1363 bp, were obtained utilising UBC889 (five polymorphic, four monomorphic, and two unique bands found at 1349 bp for Giza 177 and 651 bp for Sakha 108).Therefore, 50% of the population was polymorphic.Ten bands in all, spanning 303 to 1377 bp, were detected for the UBC807 primer (four polymorphic, six monomorphic, and 40% polymorphic).While being missing at all concentrations of ZnO NPs and MgONPs, two distinct bands with molecular sizes of 341 bp for the Sakha super 300 and 1130 bp for the Sakha 108 were found.A 42.86% polymorphism rate was present.Seven bands in all, ranging in size from 478 to 1363 bp, were obtained utilising UBC889 (five polymorphic, four monomorphic, and two unique bands found at 1349 bp for Giza 177 and 651 bp for Sakha 108).To ascertain the evolutionary relationships between the several species of barley, genetic diversity was examined in this case using the Inter-simple sequence repeat

RAPD markers:
The SBSC-18 primer produced six bands with a 33.33% polymorphism rate.Four monomorphic bands and two polymorphic bands, ranging in size from 490 to 1320 bp, were discovered in this profile.For the Sakha 108, a single distinct band with a molecular size of 977 bp was found; however, it was not present at any concentration of ZnO NPs or MgONPs.The findings collected revealed that the level of polymorphism in the primers SBSF-14 and ranged from 429 to 1511 bp, while showing five monomorphic and five polympphic bands as well as two unique bands; found at molecular sizes of 890 bp and 535 bp for the Giza 177 (Table 1&3 and Figure 7).
The two most used techniques for determining genetic differentiation across and within plant populations are RAPD and ISSR (3,4).Using RAPD primers, more polymorphic loci (56.68%) were

Genetic diversity and relationships
Seven ISSR and three RAPD markers from three different rice species were clustered using the UPGMA method, resulting in a dendrogram (Fig. 8).There were two clusters with high genetic similarity; Cluster A had one genotype that was subdivided using Sakha super 300.At a phylogenetic distance, Sakha 108 and Giza 177 were found to be clustered together.
The phenogram revealed two different clusters with genetic similarity scores ranging from 0.90 to 1.00.The Sakha super 300 and Giza 177 genotypes had a maximum genetic similarity coefficient of 0.95, indicating a high degree of genetic closeness.As shown in Table 4, the accessions Sakha 108 and Giza 177, both of which were towed, had the lowest closeness ratio of 0.90%.
Wheat has also been classified as following to this pattern of geographically linked grouping using  ).This work unequivocally showed that ZnO NPs might be an effective fertiliser and antifungal agent against pathogenic fungus that can aid in preventing M. oryzae crop infestation.In the case of the rice cultivars Giza 177, Sakha super 300, and Sakha 108, using seven ISSR primers and three RAPD primers with 24 unique bands, the obtained results using molecular markers confirmed that the lower concentrations of ZnO-NPs and MgO nanoparticles (10 and 20 mg/L) are considered to be a good enhancement agent.These findings generally indicated that lower ZnO and MgO NP concentrations might be used as an efficient nanofertilizer for sustainable agriculture and food safety, as well as an antifungal agent for the rice fungus Fusarium moniliforme (Elamawi et al., 2019).

Conclusion
When applied at the right concentration and timing, foliar application of ZnO and MgO nanoparticles on rice cultivars (Giza 177, Sakha super 300, and Sakha 108) offers a practical and useful approach to enhance rice plant performance and productivity while mitigating leaf blast.
al. (2009), Elamawi and El Shafey (2013), and Elamawi et al. (2018), silver nanoparticles are used successfully to reduce rice blast and avoid harmful infections in rice.ZnO NPs have been shown to have antifungal activity against a variety of fungus species by a number of researchers, including He et al. (2011), Gunalan et al. (2012), Wani and Shah (2012), Savi et al. (2013), Elamawi et al. (2016), and Sierra Fernandez et al. (2017).As far as we are aware, there have been no reports of ZnO NPs being used to control M. oryzae.

Fig. 1 .
Fig. 1.Zn-caNPs Characterization Dynamic Light Scattering, UV-Vis Spectra Analysis, XRD, and histogram indicate their impact on growth decrease of seed-borne fungi Fusariummoniliforme in rice that causes foot rot, along with other nanoparticle treatments.

Fig. 2 .
Fig. 2. TEM of ZnO-Zs-NPs(A) and MgO NPs (MgOac) Band MgONPs(MgOsu) C Fourier Transform Infrared Spectroscopy (FT-IR) Figure (3) shows the FTIR results for nanoparticles made from sulphate or acetate, such as ZnNPs or MgONPs.The FT-IR analysis was used to pinpoint the substances and/or functional groups that are most likely present in the created nanoparticles. the ZnONP spectrum shown in Fig. 3A, which exhibits distinct peaks at 3188.67, 2345.33,2153.2093,1959.1613.141.406.605.and 453.67 cm-1.The O-H stretching of carboxylic acid is what causes the significant broad absorption peak at 3188.67cm-1.While the peak at 1613cm-1 is attributed to C=O stretching in amides (tertiary amides) or N-H bend from amides (primary amide), possibly similar to amino acids or protein, the peak at 1063cm-1, from 605 to 429, suggests the ZnO.Potential secondary metabolites include tannins, saponins, terpinoids, and terpenes.The distinct peaks at 2103, 1437, 984, and 441 with precursor MgOac and 984cm-1 with MgOsu suggest the MgO, in addition to the emergence of the MgONPs synthesised from sulphate or acetate (MgOsu and MgOac) displayed in Fig.3B.

Fig. 5 .
Fig. 5. Agarose gel profile of genomic DNA of Rice using ISSR-PCR markers of four primers namely (UBC809, UBC823, UBC807 and UBC889).Lane numbers represent serial number of control and different concentration (mg/L) of ZnO NPs and MgONPs.M= molecular marker ladder (1500 bp).

Fig. 6 .
Fig.6.Agarose gel profile of genomic DNA of Rice using ISSR-PCR markers of three primers namely (UBC834, UBC817 and UBC873).Lane numbers represent serial number of control and different concentration (mg/L) of ZnO NPs and MgONPs.M= molecular marker ladder (1500 bp).

Table 2 .
List of ISSR primers and positive markers of three rice varieties Primer code MS Giza 177 Sakha super 300 Sakha 108 Positive markers (P+*)

Table 3 .
List of RAPD primers and positive markers of three rice varieties Primer code MS Giza 177 Sakha super 300 Sakha 108 Positive markers (P+*)

Table ( 1
): ISSR and RAPD primers used for three rice amplification, the number of bands produced, unique bands and polymorphic band percentage

Table 4 .
Elamawi et al., 2016nElamawi et al., 2016)deum vulgare L. based on 6 qualitative morphological traits.Recently, there was discussion over ZnO's ability to inhibit the growth of pathogens.Ag is probably not the best option for controlling fungal pathogens; ZnO NPs are(Dimkpa et al., 2013).For instance, ZnO NPs showed greater in vitro suppression of Helmenthosporium oryzae and F. graminearum(Dimkpa et al., 2013;Elamawi et al., 2016).Biosynthesized ZnO NPs (25 g/mL) showed greater inhibition rates against harmful bacteria and fungal species in a related investigation (Jayaseelan et al., 2012).In a plating assay, ZnO NPs considerably slowed the growth of B. cinerea and P. expansum mycelia(He et al., 2011).They mentioned that these inhibitory effects interfered with cell function, led to fungal hyphal distortion in B. cinerea, and prevented the growth of conidia and conidiophores in P. expansum.The antibacterial action of ZnO NPs has been explained by a number of different mechanisms.It is thought that the production of hydrogen peroxide from the surface of ZnO NPs is an efficient method for preventing the growth of fungi.The release of Zn2+ ions, which can harm cell membranes and interact with intracellular components, is another potential mechanism (Sirelkhatim et al., 2015).According to Navale et al.(2015), ZnO NPs and their photo catalytic characteristics significantly contribute to their antibacterial action, inducing microbial cell membrane structural alterations, oxidative stress, and ultimately the death of the cells.Contrary to the current findings,Savi et al. (2013)state that some fungi may produce more conidia as a result of the stress brought on by ZnO NPs treatments.Fortunately, research has revealed that MgO NPs cause tomato plants to activate the salicylic acid, jasmonate, and ethylene signalling pathways(Imada et al. 2016).The good effect of MgO NPs may be attributed to their indirect enhancing effect on the action of salicylic acid, which in turn boosted the rate of cell division and accumulation of IAA in wheat seedlings(Shakirova et al., 2003).Numerous studies have been done on how adding Zn to rice crops can boost grain yield in either normal or salinity soil (Swamy et al., 2016;Elamawi et al., 2016 On rice cultivars iza 177, Sakha super 300, and Sakha 108, molecular markers are regarded as useful instruments and proof of nline at: Available online at: https://jazindia.com-948 -50.Kekec, G., Sakcali, M. S. and Uzonur, I. 2010.Assessment of genotoxic effects of boron on wheat (Triticum aestivum L.) and Bean (Phaseolus vulgaris L.) by using RAPD analysis.Bull.Environ.Contam.Toxicol., 84: 759-764.51.Strelchenko P, O Kovalyova, K Okuno.(1999)."Genetic differentiation and geographical distribution of barley germplasm based on RAPD markers."Genetic resources and crop evolution; 46: 193-205.52.Owuor ED, A Beharav, T Fahima, VM Kirzhner, AB Korol, E Nevo, (2003).Microscale ecological stress causes RAPD molecular selection in wild barley, Neve Yaar microsite, Israel, Genetic Resources and Crop Evolution; 50: 213-224 53.Mahjoub A, MS El Gharbi, K Mguis, M El Gazzah, NB Brahim, (2009).Evaluation of genetic diversity in Aegilops geniculata Roth accessions using morphological and RAPD markers, Pakistan journal of biological sciences: PJBS; 12: 994-1003 54.Dakir EH, ML Ruiz, P García, MP de la Vega, (2002).Genetic variability evaluation in a Moroccan collection of barley, Hordeum vulgare L., by means of storage proteins and RAPDs, Genetic Resources and Crop Evolution; 49: 619-631