| Seed germination and seedling growth of maize (Zea mays L.) are susceptible to injury by low temperature stress. It is important to improve chilling tolerance of maize seedlings. Polyamines are small aliphatic polycationic nitrogenous compounds ubiquitous in higher plants. They are all involved in the response of plants to environmental stress. In present research, chilling-tolerant (HuangC) and chilling-sensitive (Mo 17) maize inbred lines were used to investigate the changes of the activities of catalase (CAT) and peroxidase (POD), proline (Pro) and polyamine concentration in different segments of seedlings and their relationship to chilling tolerance of maize under low temperature stress (5℃ 3 d);optimum condition on improving maize seed vigor by Put soaking was studied by employing the method of response surface analysis (RSA) under low temperature stress;relationship between polyamines, ABA and cellular membrane stability was investigated by appling exogenous polyamine, ABA with different concentrations;changes in polyamine concentration in different segments of maize seedlings, the main biosynthesis pathways of putrescine (Put) and the possibility of conversion of spermidine (Spd) to Put were investigated by applying polyamine biosynthesis inhibitors, D-arginine (D-Arg), difluoromethylormithine (DFMO), methylglyoxyl-bis- (guanyhydrazone) (MGBG) and aminoguanidine (Ag) under low temperature stress;and the genetic trait and heterosis of hybrid F1 of two maize inbred lines differing in chilling tolerance were researched. Main results were as follows:The relative growth rate (RGR) in the mesocotyl and coleoptile of HuangC significantly higher than that of Mo 17, the change rates of CAT and POD activities and Pro concentration in the three segments (root, mesocotyl and coleoptile ) of HuangC seedlings were higher than those in Mo 17 during low temperature stress (5℃, 3 d). Compared to seedlings at normal temperature, the activities of CAT and Pro concentrations in the other segments of chilled seedlings were increased except in the root of Mo 17, and the increasing extents of CAT activity and Pro concentrations in HuangC were higher than those in Mo 17. However, the POD activity in the three segments of chilled seedlings were reduced, and the decreasing extents of POD activity in HuangC was lower than those in Mo 17. The results of stepwise regression analysis showed that the accumulation of dry weights in chilling-tolerant line HuangC was mainly related to Pro concentration, however, in chilling-sensitive line Mo 17 was mainly related to POD activity during low temperature stress. The change rate of Pro concentration in mesocotyl was more closely correlation with chilling tolerance of maize inbred lines than the other physiological parameters in the other two segments.Membrane permeability (as measured by relative electrolyte (RE)), malondialdehyde (MDA) concentration and Put concentration in the three segments of the maize seedlings increased after low temperature stress (5℃, 3 d), except for Put concentration in root. However, Spd and Spermine (Spm) concentrations in the three seedling segments decreased after chilling stress. A relative change level in both Put and Spd, and the ratio of Spd to total polyamines in HuangC was higher than those of Mo 17, as compared different segments, coleoptile was higher than mesocotyl in both inbred lines. However, for chilling injury parameters, HuangC was lower than Mo 17, comparing different segments, coleoptile was lower than mesocotyl in both inbred lines. The results of regression analysis indicated that chilling injuryparameters in root were mainly correlated with Spd concentration, in mesocotyl were mainly with Put and Spd concentration and in coleoptile were mainly with Spd and Spm concentration. Therefore, the characteristics of polyamine changes in chilling-tolerant maize seedling combining with regression analysis will be more reliable for evaluating relative chilling tolerance among maize inbred lines.Two factors (concentrations of Put and soaking time) with 14 combinations of RSA experiment method was designed to treat chilling-sensitive Mo 17 seeds, then the low temperature germination experiment was carried out. After root and shoot dry weight, RGR and RE were analyzed by RSA, the optimum combinations of Put treatment for increasing shoot dry weight, root dry weight, shoot RGR and decreasing shoot RE was found. According to these optimum combinations, the optimum ranges for Put soaking concentration and soaking time were 0.45-0.60mM and 11.5-17.9 h.The effects treated seedlings by Put> Spd and Spm (O.lmM, 0.5mM, l.OmM, 2.0mM ) and abscisic acid (ABA) (5uM, lOuM, 50uM, lOOuM) before low temperature stress on seedling growth, membrane stability (indicator by MDA) and Chlorophyll (Chi) contents were investigated. The results indicated that low concentrations of 0.5mM Put, O.lmM Spd and O.lmM Spm treatments and 5uM ABA on the improving seedling growth, reducing MDA concentrations and increasing Chi contens had better effects. Among Put, Spd and Spm treatments, the most effective treatment was Spd, followed was Put, the worst was Spm. High concentrations of polyamine (2.0mM) and ABA (50uM, lOOuM) treatments were negative effects. It indicated that the relationship between polyamine and cellular membrane stability, the improvement of chilling tolerance of maize seedlings were all correlated with polyamines and ABA concentrations and components of polyamines.MGBG treatment reduced the Spd concentration, increased membrane permeability and MDA concentrations in three segments of both maize inbred lines seedlings before and after low temperature treatment (5°C, 3 d), and decreased the length and fresh weight of root and shoot, chl contents at the end of recovery growth. It suggested that MGBG treatment increased the maize seedling chilling injury. Put or Spd plus MGBG treatment alleviated the lipid peroxidation (indicator by MDA concentration), improved seedlings growth and increased Chl contents comparing to MGBG treatment, however, Spd plus MGBG was more effective than that of Put plus MGBG It suggested that Spd may be play an more important role in chilling tolerance.D-Arg and DFMO treatment inhibited the biosynthesis of Put in three segments of both inbred lines after suffering low temperature stress suggesting the Put biosynthesis may be affected by both ADC and ODC pathways. However, when maize seedlings treated by the same concentration of D-Arg or DFMO, the Put concentration in seedlings by DFMO treatment was significantly lower than that by D-Arg treatment. It indicated that biosynthesis of Put was mainly via ODC pathway, minorly via ADC pathway. Low concentration (10uM) of D-Arg and DFMO improved seedling growth, increased the length and fresh weight of root and shoot and Chl content of seedling leaves under low temperature, moreover, the effects of D-Arg treatment was better than that of DFMO treatment. It may be related to that DFMO inhibited Put synthesis more effective than D-Arg.Combinations of four polyamine inhibitors, D-Arg and DFMO (inhibiting the biosynthesis of Put), Ag (inhibiting the degradation of Put) and MGBG (inhibiting the biosynthesis of Spd and Spm from Put) with different concentration ((1) lOuM combinations included lOuM D-Arg, lOuM DFMO, lOuM MGBG, lOOuM Ag;(2) lOOuM combinations included lOOuM D-Arg, lOOuM DFMO, lOOuM MGBG, lOOOuM Ag;(3) 500uM combinations included 500uM D-Arg, 200uM DFMO, 500uM MGBG, 5mM Ag;(4) lOOOuM combinations included lOOOuM D-Arg, 200uM DFMO, lOOOuM MGBG, lOmM Ag) were applied to investigate whether Spd can convert to Put or not. Combination treatments of polyamineinhibitors inhibited the biosynthesis of Put in three segments of both maize inbred lines. There was no significant difference in Put concentration between 500jtM and 1000/iM polyamine inhibitor combinations except for Mo 17 coleoptile. Therefore, Spd plus 1000/iM polyamine inhibitor combination was selected to treat maize seedlings sequentially, Put concentration in the treated root was 2.54-5.23 times, in mesoctyl was 1.34-1.48 times, in coleoptile was 1.47-1.49 times compared to the control of 1000/tM combination, respectively. It indicated that Spd may be converted to Put, moreover, this conversion in root was stronger than that in mesocotyl and colepoptile.Heterosis of hybrid Fl of two maize inbred lines differing in chilling tolerance was researched under low temperature stress. Length and fresh weight of root and shoot of the Fi seedlings were higher than those in the parents. Chi a, carotenoid and total Chi content in the Fi were all higher than those in the parents. After maize seedling growth for 4 days at normal temperature, Spd and total polyamine concentration in the three segments of the Fi seedlings had significant heterosis in heterosis of mid-parent, heterosis of over high-parent, and index of heterosis, however, Put and Spm no heterosis, total ployamine had heterosis in heterosis of mid-parent and index of heterosis. After suffering low temperature stress for 3 days, Spd and total polyamines concentrations in three segments of the Fi seedlings, except for total polyamines concentrations in coleoptile, heterosis in other segements was simlar to at normal temperature. After recovery growth at normal temperature for 3days, Spd and toatl polyamine concentration lost heterosis, only Put in mesoctyl had heterosis. MDA had a strong negative heterosis, sugesting that the hybrid Fj from chilling-tolerant crossed chilling-sensitive inbred lines reduced lipid peroxidtion may be related to the increase of Spd concentration in different segments of the Fi seedlings.
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