Bazı buğday (Triticum aestivum L.) genotiplerinde nikel toksisitesi etkilerinin çimlenme ve erken fide evresinde fizyolojik olarak araştırılması
Physiological investigation of the effects of nickel toxicity in some wheat (Triticum aestivum L.) genotypes at the germination and early seedling stage
Yazar: TUĞBA ÖZDEMİR
Danışman: DR. ÖĞR. ÜYESİ ALİ DOĞRU
Sakarya Üniversitesi / Fen Bilimleri Enstitüsü / Biyoloji Ana Bilim Dalı
Derece: Yüksek Lisans, 2024
Konu: Biyoloji = Biology

Özet:
Bu çalışmada buğday (Triticum aestivum L.) bitkisine ait yedi farklı genotipin (Cumhuriyet-75, Efe, Gönen-98, Kaşifbey-95, Kayra, Meltem ve Ziyabey-98) çimlenme ve erken fide evresinde nikel toleransları araştırılmıştır. Çalışma iki aşamada gerçekleştirilmiştir.
Birinci aşamada kurulan denemede buğday genotipine çimlenme aşamasında farklı konsantrasyonlarda (0, 200, 400, 600, 800 ve 100 ppm) nikel (NiSO4 .7H2O) çözeltileri uygulanmış, denemenin sekizinci gününde hasat yapılarak bazı çimlenme ve fizyolojik büyüme parametreleri ölçülmüştür. Buna göre fidelerin kök, koleoptil ve toplam boyu; kök ve koleoptillerin taze ve kuru ağırlığı; kök ve koleoptillerin gerçek ve oransal büyüme hızı; tohumlardaki kullanılan depo maddelerinin miktarı, konversiyon kat sayısı ve depo maddelerinin tüketimi (%) uygulanan nikel konsantrasyonuna bağlı olarak azalmıştır. Kök ve koleoptil boyları esas alınarak hesaplanan tolerans indekslerine göre yapılan puanlama sonucunda çimlenme evresinde nikel toksisitesine en dayanıklı genotipin Ziyabey-98, en duyarlı genotipin ise Cumhuriyet-75 olduğu belirlenmiştir.
Çalışmanın ikinci aşamasında Cumhuriyet-75 ve Ziyabey-98 genotipleri kullanılarak saksı denemeleri kurulmuş ve 200 ppm nikel uygulaması yapılmıştır. Nikel toksisitesi Ziyabey-98 genotipinde kök büyümesinin, Cumhuriyet-75 genotipinde ise gövde büyümesinin ilgili kontrollere göre önemli derecede inhibe olmasına yol açmıştır. Bu sonuç her iki genotipin kök ve gövde büyümesinin nikel toksisitesine farklı derecede toleransa sahip olduğunu göstermiştir. Kök ve gövdenin taze ve kuru ağırlıklarında nikel toksisitesi etkisiyle meydana gelen değişimler, her iki genotipte de gövdenin daha duyarlı olduğunu göstermiştir. Toplam SOD aktivitesi nikel uygulanan Ziyabey-98 genotipinde artmış, Cumhuriyet-75 genotipinde değişmemiştir. Bu durum muhtemelen Ziyabey-98 genotipinde nikel toksisitesinin yaprak dokularındaki süperoksit birikiminin artmasından kaynaklanmıştır. Toplam APOD aktivitesi nikel uygulaması sonucunda Cumhuriyet-75 genotipinde azalmış, Ziyabey-98 genotipinde artmıştır. Toplam GR aktivitesi ise nikel toksisitesi altındaki Cumhuriyet-75 genotipinde değişmemiş, Ziyabey-98 genotipinde artmıştır. Bu sonuçlar nikel uygulanan  Ziyabey-98 genotipinde askorbat-glutatyon döngüsünün aktif olduğunu ve etkili bir şekilde H2O2 detoksifikasyonunun gerçekleştiğini göstermektedir. Toplam GPOD aktivitesi ise nikel toksisitesi altındaki her iki genotipte de değişmemiştir. Ölçülen klorofil a floresansı ve hesaplanan JIP testi parametrelerinde gözlenen değişimler, Cumhuriyet-75 genotipinde fotosentetik birimlerin ve elektron taşınım reaksiyonlarının nikel toksisitesinden etkilenmediğini; Ziyabey-98 genotipinin ise olumsuz yönde etkilendiğini göstermiştir.
Buna göre her iki genotipin çimlenme ve erken fide evresinde nikel toksisitesine karşı tolerans ve duyarlılık derecelerinin farklı olduğu, erken fide evresinde uygulanan nikel toksisitesinin Ziyabey-98 genotipinde fotosentetik elektron taşınım reaksiyonlarını bozarak oksidatif strese neden olduğu söylenebilir

Abstract:
In this research, the growth parameters and some physiological and biochemical changes caused by nickel (NiSO4.7H2O) stress on the germination and seedling stages of the wheat (Triticum aestivum L.) plant were studied.
In the first stage, different concentrations of nickel (200, 400, 600, 800 and 1000 ppm) solution were used to apply on seven different genotypes of wheat plants (Cumhuriyet-75, Efe, Kayra, Gönen-98, Kaşifbey-95, Meltem and Ziyabey-98). has been prepared. The same number of seeds and the same size were placed in the containers between filter papers moistened with their own solution, in five replicates. On the third day, 1 mM intermediate irrigation was applied with its own concentrations, and on the eighth day it was harvested and relevant analysis and measurements were made.
As a result of nickel application, it was determined that there was a statistical decrease in total root and stem length compared to the control in all genotypes (p<0.05). A statistically significant decrease in root fresh weight was observed at concentrations of 400 ppm and above for all genotypes (p<0.05). When the root dry weight was examined, the most resistant genotype was Kaşifbey-95, with no significant change (p>0.05).
When the stem fresh weights are compared to the control, significant inhibition is observed at concentrations of 600 ppm and above for all genotypes (p<0.05). Kaşifbey-95 was the most affected by the decrease in hull dry weight due to nickel application (p<0.05).
All nickel (NiSO4.7H2O) concentrations applied to Efe and Ziyabey-98 genotypes caused a significant decrease in the actual growth rate in the root (p<0.05). All nickel (NiSO4.7H2O) concentrations (200, 400, 600, 800 and 1000 ppm) applied on Gönen-98, Cumhuriyet-75, Ziyabey-98 genotypes caused a significant change in the decrease in the actual growth rate of the trunk compared to the control group (p <0.05). The relative growth rate of root and stem was significantly inhibited in Efe, Kayra, Gönen-98, Meltem and Ziyabey-98 genotypes at concentrations of 600 ppm and above, and this was statistically significant (p<0.05). 
Gönen-98 ve Ziyabey-98 genotipleri 200 ppm ve üzeri konsantrasyonlarda uygulanan nikel kontrole göre istatistiki açıdan önemli olan kullanılan rezerv miktarında azalmaya sebep olmuştur (p<0.05). Statistically, nickel application of 200 ppm and above caused a significant decrease in Kaşifbey-95 and Kayra genotypes. Nickel application significantly reduced the reserve consumption (%) rate of the Meltem genotype at all concentrations (p<0.05).
When compared among the wheat genotypes studied, it was concluded that the most damaged and most sensitive genotype against nickel toxicity was Cumhuriyet-75, and the least damaged and most resistant genotype was Ziyabey-98.
In the second stage, approximately the same size and sound seeds of two genotypes of wheat (Triticum aestivum L.) genotypes, Cumhuriyet-75 and Ziyabey-98, were selected. Seeds were placed between filter papers moistened with bidistilled water in lidded containers. The containers were left to germinate in a climate cabinet at 24 oC and 40-50% relative humidity. Three days later, uniform seedlings of both genotypes were transferred to pots containing perlite and ½ diluted Hoagland nutrient solution and maintained at 18/25 oC temperature (day/night), 16/8 hour photoperiod (day/night), 50±5% relative humidity and 200 µmol photons were placed in a climate cabinet with a light intensity of m-2s-1. Twelve day old plants are divided into 2 groups. 
The control plants in the first group were watered with ½ diluted Hoagland nutrient solution until the end of the experiment, the plants in the second group were given 200 ppm nickel (NiSO4.7H2O) solution, two days later, photosynthetic activity was measured on the leaves of the plants with the chlorophyll a fluorescence technique, some physiological growth parameters. Leaf samples to be used in biochemical analyzes were stored at -20 °C until analysis. 
When the data obtained at the seedling stage are examined, 200 ppm nickel application causes a significant change in root length in the Cumhuriyet-75 genotype compared to the control group, while a significant decrease in stem length is observed in the Ziyabey-98 genotype (p<0.05). Application of 200 ppm nickel did not cause a statistically significant decrease in total height in both Cumhuriyet-75 and Ziyabey-98 genotypes (p>0.05).Nickel (200 ppm) application caused a significant decrease in root fresh weight, stem fresh weight and stem dry weight in Ziyabey-98 genotype (p>0.05).  However, the change in root dry weight in Cumhuriyet-75 and Gönen-98 genotypes was not significantly affected by nickel (p>0.05). Wheat genotypes have created some reactions with defense mechanisms against nickel stress. These reactions are carried out by enzymatic and non-enzymatic means. In response to 200 ppm nickel application, total superoxide dismutase (SOD) enzyme activity increased significantly in Ziyabey-98 compared to its own control (p<0.05), while the SOD increase in the application to Cumhuriyet-75 genotype was statistically insignificant (P>0.05). Total SOD activity was calculated using the standard graph (U/mg protein) according to Beyer and Fridovich (1987). Total APOD activity Wang et al. According to (1991); It was calculated from the initial rate of the reaction (nmol ascorbate min-1 µg protein-1) using the extinction coefficient of ascorbate (2.8 mM cm 290 nm-1). While 200 ppm nickel application causes a significant increase in APOD activity in the Cumhuriyet-75 genotype, it causes a statistically significant decrease in APOD activity in the Ziyabey-98 genotype (p<0.05). While the total Glutathione peroxidase (GR) enzyme activity caused its own control and a statistically significant increase against the applied nickel, it did not affect the Cumhuriyet-75 genotype. The total GR activity was determined by Sgherri et al. according to (1994), it was calculated from the initial rate of reaction using the extinction coefficient of NADPH (6.2 mM cm 340 nm-1) (nmol NADPH per minute -1 µg protein-1).
GOOD activity caused an increase in Cumhuriyet-75 genotype and a decrease in enzyme activity in Ziyabey-98 genotype, and these changes were statistically significant (p<0.05). Total POD activity was calculated from the initiation rate of the reaction (nmol H2O2 min -1 µg protein-1) using the extinction coefficient of guaiacol (26.6 mM cm 470 nm-1) according to Sanchez-Romero (1993).
Chlorophyll a fluorescence parameters were determined in the leaves with a "plant productivity analyzer" (HandyPEA fluorometer, Hansatech Instruments Ltd., Pentney, King's Lynn, Norfolk, England), and the obtained data were evaluated according to the JİP test.
The results of  200 ppm nickel application in Cumhuriyet-75 genotype compared to the control, Fo (minimum fluorescence), Fm (maximum fluorescence), tFm, Fv/Fm ratio (maximum quantum efficiency of FS II), Fv/Fo (Hill reaction efficiency), V/to (accumulation rate of closed reaction centers) Area (region above the OJIP curve and between Fo and Fm) Po, o, Eo, Do, Ro, Ro, ABS/RC, TRo/RC and ETo/RC, DIo/RC (dissipation energy lost excluding photochemical events per reaction center in FS II; at t=0), SFI(abs) (indicator of the structural and functional state of FS II), RC/ABS (FS amount of active reaction centers per antenna chlorophylls in II), o/(1-o) (Performance indicator of photochemical reactions), o/(1-o) (Performance indicator of non-light-dependent (dark) reactions) and PI Changes in (abs) (Performance index) values are statistically insignificant (p>0.05). Therefore, 200 ppm nickel application did not affect the photosynthetic activity in Cumhuriyet-75 genotype.
Ziyabey-98 genotype, the change in tFm value, decrease in Fm value, increase in PI (abs) and increase in Fo do not cause a statistically significant change when compared to the control group with 200 ppm nickel stress (p<0.05). In Ziyabey-98 genotype,Fv/Fm, Fv/Fo, Area, Po, o, Ro, Eo, SFI(abs), RC/ABS, o/(1-o), o/(1-o) values and the increase in V/to, Do, Ro, ABS/RC, TRo/RC, ETo/RC, DIo/RC values are statistically significant (p<0.05). Application of 200 ppm nickel reduces the photosynthetic activity of Ziyabey-98 genotype.
As a result of the studies, it can be concluded that nickel stress affects the physiological and biochemical activities of the wheat plant, increases some enzyme activities and decreases some enzyme activities, and generally has a negative effect on growth parameters and photosynthetic activities. The reason why it leaves different effects is because the wheat has different genotypes. There are differences in the resistance and sensitivity rates of genotypes to applied stress.