| Foxtail millet has become an important food crop in northern China because of its strong resistance and high nutritional value.In order to improve the differentiation rate of foxtail millet callus regeneration system established in the previous experiment,Jingu 21 foxtail millet seeds were used as explants to the effects of adding different kinds and concentrations of hormones,concentrations of sucrose,adding additional substances and subculture on the callus induction.A three-factor and five-level orthogonal experiment was designed to study the effects of sucrose concentration,growth retarder concentration,and drying treatment on the differentiation of callus in order to establish an efficient and stable foxtail millet callus regeneration system.At the same time,Jingu 21,Yugu 35 and Jigu 42,which have different resistance to sethoxydim,were used as materials,and different concentration of sethoxydim was added to the medium,to understand their effects on seed germination,callus induction and growth of three foxtail millet cultivars,and the callus of three foxtail millet cultivars under sethoxydim stress were used as materials,15 physiological and biochemical indexes were measured,and the physiological response and resistance mechanism of three foxtail millet cultivars to sethoxydim stress were studied from the cell level.This experiment provides a theoretical basis for the application of genetic transformation in foxtail millet and the further study of herbicide resistance in foxtail millet.The results are as follows:1.Jingu 21 foxtail millet seeds were used as explants to induce embryogenic callus which was easy to redifferentiate.The results showed that the effect of adding 6-Benzylaminopurine(6-BA)and2,4-Dichlorophenoxyacetic acid(2,4-D)in the medium was better than that of kinetin(KT)and thidiazuron(TDZ);adding a small amount of abscisic acid(ABA)and silver nitrate(Ag NO3)could improve the callus induction rate;properly increasing the sucrose concentration in the medium could improve the callus induction rate and the quality of induced callus,which was good for differentiation.Therefore,the most suitable medium for callus induction was MS + 2.0 mg/L 2,4-D + 0.2 mg/L 6-BA + 5 % sucrose.2.A three-factor and five-level orthogonal experiment was designed to study the effects of sucrose concentration,growth retardants concentration in the medium and drying treatment on callus differentiation.The results showed that the differentiation rate of callus could be improved by properly reducing the concentration of sucrose and increasing the concentration of agar,and adding a small amount of PP333.The quality of differentiated plantlets was better than that of the control.The suitable medium for callus differentiation was MS + 1.0 mg/L 6-BA + 0.5 mg/L NAA + 2.5 % sucrose + 14 g agar + 4 mg/L PP333,MS+ 1.0 mg/L 6-BA + 0.5 mg/L NAA + 3 % sucrose + 14 g agar + 3 mg/L PP333,MS + 1.0 mg/L 6-BA + 0.5mg/L NAA + 2 % sucrose + 10 g agar + 4 mg/L PP333.In these three mediums,the callus differentiation rate is high and the differentiated seedlings are relatively robust.3.We took the seeds of Jingu 21(sensitive to sethoxydim),Yugu 35 and Jigu 42(resistant to sethoxydim)as materials,the effects of sethoxydim on seed germination,callus induction were observed.We took the callus of three kinds of millet,observed the effects of sethoxydim on callus growth,and measured 15 physiological and biochemical indexes such as malondialdehyde(MDA),superoxide anion(O2-)content,membrane permeability(MP),superoxide dismutase(SOD),peroxidase(POD),catalase(CAT),aseorbateperoxidase(APX),glutathione reductase(GR)activity,proline(Pro),soluble protein(SP),soluble sugar(SS),ascorbic acid(As A),docosahexaenoic acid(DHA),As A+DHA,glutathione(GSH)content,etc.Correlation analysis were carried out among different indexes of the same variety and different varieties.The results showed that the increase of sethoxydim concentration had a significant effect on seed germination,callus induction and callus growth of three foxtail millet cultivars,and the effect on Jingu 21 foxtail millet was significantly greater than Yugu 35 and Jigu 42.Sethoxydim treatment increased O2-accumulation and MDA content in callus cells of three foxtail millet cultivars,and increased MP.We analyzed the activity the changes of antioxidant enzymes SOD,POD,CAT,GR,APX,the changes in the content of the osmotic regulation substances Pro,SP,SS,and antioxidants As A,DHA,and GSH between sensitive and resistant varieties.It was found that the change of these indexes of resistant varieties Yugu 35 and Jigu 42 was significantly different from susceptible varieties Jingu 21,and the change of these indexes was good for resisting sethoxydim stress.At the same time,the changes of the above indexes were different between the two resistant varieties Yugu 35 and Jigu 42,which indicated that the resistance mechanism of foxtail millet herbicide was complex and was the result of comprehensive defense in different aspects. |
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