Cold resistance evaluation of the shoots of 5 apple rootstocks

 Abstract:【Objective】The cold resistance of the hardwoods of different root-stocks was evaluated com- prehensively in order to select rootstocks that improve the cold resistance of‘Nagano Fuji No.2’apple, which is susceptible to cold injury in winter. Cold injure usually causes yield reduction and even crop failure, which is disastrous for farmers and seriously affects the development of apple industry. The ex- isting studies mainly focus on frozen injury in apple in winter, but few reports concern about the mecha- nisms of response to cold temperatures. In this study, we used the membership function and the semi-le- thal temperature(LT50)to analyze changes in antioxidant enzymes and osmotic regulators in apple under cold stress in order to provide a theoretical reference for cultivar introduction, cold resistance breeding and cultivation【. Methods】One-year-old hardwoods of five apple rootstocks including SH1, Y-1, B9, T337 and M26 were placed in incubators for low temperature treatment. The experiment set six temper- ature treatments including exposure of the hardwoods to room temperature (CK), -15 °C, -25 °C,-30 °C, -35 °C and -40 °C under darkness for 12 h. Each treatment composed of three replicates. Sam- ples were taken to determine the membrane leakage reflected by relative electric conductivity (REC), or frozen in liquid nitrogen then stored at -80 °C for analyses of malondialdehyde (MDA), contents of sol- uble sugars and free proline and the enzymatic activities of peroxidase (POD) and superoxide dismutase (SOD).【Results】The REC value of the hardwoods of the five cultivars increased following an S curve   with the decrease in temperature. The rapid increase in REC in different cultivars occurred at different temperatures. The REC values of T337 and M26 increased significantly when temperature was be- low -25 °C; SH1 exposed to cold stress for 12 h behaved similar to Y-1. When temperature was lower than -25 °C, the REC values of SH1 and Y-1 significantly rose. Regressed logistic equations between temperature and REC values in the five cultivars exposed to the cold temperatures for 12 h were estab- lished. LT50 value was calculated from the established equations, which were used to measure the cold resistance of apple. The cold resistance based on LT50 in the five rootstock cultivars was in the order of B9 > SH1 > Y-1 > M26 >T337. MDA content in all the cultivars was higher than in the control. MDA content in all the cultivars gradually increased with the decrease in temperature, and reached the highest value at -35 °C. The highest MDA was found in SH1 and the lowest in Y-1. MDA in T337 was higher than in the other cultivars. B9 had the lowest MDA content among all the cultivars. The range of in- crease in MDA was the largest in T337 and lowest in B9 and SH1. As the temperature became lower, SOD and POD activities showed different change patterns in different cultivars. SOD activity in SH1 and Y- 1 rose first and then declined. We set that under low temperature stress of CK - 40 °C , SOD curves of B9 and SH1 changed similarly, both of which were“M”type. The SOD activity of SH1 showed“N”type under low temperature stress, reached the lowest peak at a certain inflection point of low temperature stress, and then continued to increase. Y-1 maintained a significantly higher SOD activity than other cultivars at all the low temperatures, followed by SH1 and B9. Low temperature stress in- creased SOD activity. Compared with the control group, the increasing range of T337 was larger, while those of SH1 and Y-1 were smaller than the others. POD activity in B9 and T337 increased in the early period and then decreased. The enzyme activity in Y-1 and SH1 displayed a trend of decreasing -increas- ing- decreasing, but its change was slight. POD activity in M26 was significantly higher than that in the other cultivars. In all the temperature treatments in M26, POD activity was lower than in the other culti- vars. POD activity in the five rootstock cultivars exposed to the low temperatures was higher than in the control. POD activity significantly rose when temperature dropped to below - 30 °C , and the rising range was the largest in T337 and smallest in M26. CAT activity was higher in cold stressed groups than in the control. SH1 and Y-1 had their peak CAT activities at -25 °C and T337 and M26 at -35 °C. Seven treatments in B9 were lowest among all the cultivars. Same to POD, the largest and lowest in- creasing range was found in T337 and M26, respectively.【Conclusion】REC, MDA content, solu- ble sugar, and SOD and POD activities were increased by cold stresses. With temperature drop, REC and MDA content in the five rootstock cultivars increased; the change patterns of SOD and POD activi- ties differed among cultivars. The LT50 and the changes in protective enzyme activities showed that cold resistance in the cultivars was in the order of B9 > SH1 > Y-1 > M26 > T337.