Evaluation of diversity of lentil (Lens culinaris Medik.) genotypes under freezing stress in controlled conditions

Lentil (Lens culinaris Medik.) is an important legume that plays a significant role in food security and human nutrition in the world. Lentils provide protein and fiber, as well as many vitamins and minerals, such as iron, zinc, folate, and magnesium. Lentil is a moderately drought tolerant crop, but the yield is drastically reduced with increased drought stress. One of the simplest ways to reduce the effects of drought stress is regulate plant growth period to avoid moisture stress; termed as drought escape; therefore, autumn planting can be effective in reducing the effects of drought stress in lentile. On the other hand, cold and freezing are the most important factor limiting lentil cultivation in autumn planting. Considering the importance of autumn planting in cold and highlands areas to use the seasonal rainfall in lentile crop and also due to the diversity among lentil genotypes for cold tolerance and the importance of lentil as a source of high nutritional value, this study was conducted to identify cold tolerant lentils genotypes.

This research was carried out in order to investigate the effective traits in freezing tolerance of lentil genotypes, as factorial based on Completely Randomized Design with three replications under controlled conditions at Ferdowsi University of Mashhad in 2020. The studied factors included 18 lentil genotypes at four freezing temperatures (0, -15, -18 and -20 °C). The pots were irrigated 24 hours before the freezing stress and then transferred to the thermogradient freezer to apply the tretments in mid-February. The freezer temperature at the beginning of the experiment was 5 °C and after placing the samples with slope of 2 °C per hour the temperature decreased. In order to create ice nucleation in the plant and to avoid the supercooling phenomenon, at 3 °C, Ice nucleation active bacteria (INAB) were sprayed on the plant. In order to balance the ambient temperature, seedlings were kept in each temperature treatment for one hour and then overnight in a cold room at 5 °C. Before exposing the plant to freezing stress, photosynthetic pigments, DPPH radical activity, anthocyanin, total phenol, soluble carbohydrates, malondialdehyde (MDA), proline content, catalase activity, peroxidase activity, and the relative water content (RWC) of the osmotic potential were measured. Three weeks after transferring the samples to the greenhouse, the survival percentage of the samples were evaluated. Plant survival percentage was calculated by counting the number of live plants before and after frost stress in each pot.

The results showed that lowering the temperature to -18 and -20°C reduced the survival rate in most genotypes. The highest survival percentage was observed in MLC11 genotype at -18°C. None of the studied genotypes could withstand temperatures of -20°C. At -15°C, MLC13, MLC17, MLC70, MLC409 and MLC454 genotypes had a survival of over 80%. Factor analysis showed that the first factor accounted for 31.12% of the changes with chlorophyll a, carotenoids, Cha to Chb ratio, total photosynthetic pigments and inhibition of DPPH free radical activity and the second factor accounted for 18.28% of the changes with chlorophyll b, peroxidase, plant height and biomass justifies. Due to these traits, MLC8, MLC13, MLC17, MLC38, MLC84, MLC286 and MLC334 genotypes are considered as high stress tolerance genotypes. Analysis of genotype clusters and comparison of group means showed that all traits except soluble carbohydrates, proline, relative leaf water content, catalase and osmotic potential in the first group (MLC8, MLC11, MLC33, MLC47, MLC70, MLC84, MLC4, MLC409, MLC409) They were superior to the total average.

Significant variations were observed among the genotypes studied in terms of survival rate, regrowth, and antioxidant traits. Clustering and mean comparison analysis revealed that genotypes in the first group exhibited superior cold tolerance. These genotypes outperformed the overall average in most of the examined traits. On the other hand, genotypes in the second and third groups had lower mean survival rates compared to the overall mean, indicating their higher sensitivity to stress. The first group included genotypes MLC8, MLC11, MLC33, MLC47, MLC70, MLC84, MLC409, MLC454, and MLC472. Further investigations of these genotypes under field conditions are recommended to explore their potential and performance.