Evaluation of salinity tolerance of different sorghum cultivars at seedling stage and its relationship with SSR molecular markers

Master's thesis in the field of agriculture, plant breeding

Abstract

D

Salinity stress, after drought stress, is the second limiting factor in the yield of agricultural plants on the surface It is universal. Based on the division of plants in terms of tolerance to salinity stress, sorghum is placed in the semi-tolerant class, and its salinity tolerance threshold is 4.9 to 6.8 decisiemens/meter. It is very sensitive in the vegetative stage and the early reproductive stages, while in the flowering period and the seed filling stage, they are the least sensitive to salinity stress, respectively. In order to identify salinity tolerant cultivars in sorghum (in the seedling stage) and to find SSR molecular markers related to salinity tolerance characteristics, this research was conducted in two separate experiments as follows. The first experiment was conducted as a factorial in the form of a completely random basic design with 3 replications. The first factor included salinity stress at 4 control levels (Hoagland's solution), 5, 10, and 20 dS/m, and the second factor included 10 sorghum genotypes (Sor834, Sor1003, Sor1009, Sor1011, Sor857, Sor808, Sor1006, MTS, LTS, and HTS). 20 days after applying salinity treatments, root dry weight, leaf dry weight, stem length, leaf surface, proline content, soluble sugar, sodium, potassium, magnesium and leaf calcium were measured. The interaction of salinity and genotype was significant at 1% level for all traits. Under salt stress conditions, there was a positive and significant phenotypic correlation at the 1% probability level between leaf dry weight and potassium content. The genetic correlation between calcium and leaf potassium was negative and significant and with proline and sugar content was positive and significant. Leaf dry weight had the most positive and significant genetic and phenotypic correlation with leaf potassium under salt stress conditions. The grouping of genotypes based on Yp, Ys and STI three-dimensional diagram showed Sor834, Sor1011, Sor1006 and Sor857 genotypes for cultivation in saline and non-saline conditions, which, considering other characteristics, it is possible to introduce Sor834 genotype as a salinity tolerant genotype in seedling stage. The results of the stress intensity showed that the trait of sodium to potassium ratio with a value of 0.96 was most affected by salinity stress. In the second experiment, 10 pairs of microsatellite primers were used to investigate the genetic diversity of sorghum genotypes. After DNA extraction and fragment amplification using SSR primers, the resulting bands were scored. And statistical analysis was done. The average polymorphic information content (PIC) for the evaluated primers was equal to 0.48, and the average observed and expected heterozygosity for all primers was calculated as 0.29 and 0.39, respectively. Cluster analysis based on molecular data using Jaccard similarity coefficient and UPGMA method placed sorghum cultivars into 4 main groups. Based on the results of decomposition into principal components, the first two components, namely PCA1 and PCA2, explained 50.08 and 10.08 respectively (totally 60.15%) of the total variance. That is, the SSR markers used have a relatively suitable distribution at the genome level. The results of binary logistic regression indicated a possible connection between ALL1-P6 and ALL2-P8 alleles with stress tolerance and leaf dry weight index traits, which in case of final confirmation, can be used to screen salinity tolerant genotypes.

Key words: logistic analysis, salinity, sorghum, SSR indicator

The importance of sorghum

Sorghum with the scientific name (Sorghum bicolor L.), is a C4 group plant, one year that belongs to the cereal family (Doggett, [1] 1970). Sorghum is a low-expectation and high-yielding plant that is cultivated even in poor lands in terms of nutrients or salt and alkali, so that in terms of endurance and contentment, it is called a plant camel (Kirimi, 1996). It is also a short-day plant, generally an annual, that has been cultivated for centuries by natives of Africa, the Near East, and the Middle East. In terms of production, sorghum ranks fifth among cereals after wheat, rice, corn and barley (Imam, 2006). Sorghum is the main grain in Africa, India, parts of China, Central and South America, Australia, Argentina and Mexico. Its grain yield is very different depending on weather conditions, so that it is mentioned between 300-3000 kg per hectare in some sources.The distribution of species related to cultivated sorghum in Africa indicates the domestication and cultivation of this crop in this continent. Sorghum is consumed as a staple food in many countries with warm climates. In addition, sorghum is used to produce starch, alcohol, sugar, dextrose and edible oil. Sorghum seeds are used as food supplements and concentrates for livestock and poultry, and its stem and leaves are also used to feed cattle, prepare building materials, and as fuel (Imam, 2006).  In general, environmental stress is one of the important factors limiting the performance of agricultural plants at the global level. Environmental stresses, by affecting the biochemical and physiological processes of the plant, reduce the yield of the agricultural plant and in severe cases even cause the death of the plant. Among the abiotic stresses, salinity stress is the second limiting factor for the yield of agricultural plants at the global level after drought stress. More than 6% of the world's land and nearly 50% of the world's water lands have salinity problems (Flor [2] et al., 1997). One of the most important agricultural problems in Iran is soil salinity. About 10% of Iran's soils are saline and sodium soils. Their occurrence may be natural, or accelerated by the development of irrigation and the intensification of water use with high evaporation rates and human activity (Abrol[3] et al., 1988). The response of plants to salinity stress is different and depends on the degree of toxicity and osmotic potential of salt and the duration of the stress (Kumba [4] et al., 1998). Salinity stress is a factor that seriously limits the production of crops in different regions, including arid and semi-arid regions. Salinity is a problem in most of the sorghum growing areas in the world (Taylor [5] et al., 1975). Based on the division of plants in terms of tolerance to salinity stress, sorghum is placed in the semi-tolerant class, and its salinity tolerance threshold is 4.9 to 6.8 decisiemens/meter. But among cultivars of a crop species, tolerance to salt stress can be very different. The effects of salinity on sorghum plant are different in different stages. Sorghum is very sensitive to salinity stress in the vegetative stage and the early reproductive stages, while it is the least sensitive to salinity stress in the flowering period and the seed filling stage, respectively. Salinity sometimes affects the timing of plant growth stages of some cereals, such that germination, spike emergence, and flowering occur earlier. This reaction has been reported for wheat, sorghum and oat plants (Mir Mohammadi Meibdi and Qarayazi, 2011). Today, molecular markers are used as a suitable and complementary tool to classical methods for locating and identifying genes controlling quantitative traits and selection programs. Identifying molecular markers connected to the desired gene and locating it on the chromosome is an important goal in plant breeding for gene homogenization and marker-assisted selection (Arif [6], 2002). The use of molecular methods can provide useful information related to the issues related to the characteristics of genetic diversity, such as recognizing and measuring the differences in genetic compositions, their degree of similarity, the degree and cause of diversity and kinship relationships between them. Diversity can be investigated at the level of species, population and within the population (individuals) (Bagheri et al., 2018).  The degree of polymorphism is one of the important characteristics of each marker, which expresses its relative importance, in addition, the usefulness of a marker is related to the number of alleles and their frequency. On the other hand, allelic frequencies indicate the genetic structure of societies. In general, any signs or symptoms that indicate the presence or absence of a trait or a specific feature in a plant are called indicators. It should be noted that every marker used in plant breeding must have two characteristics. One is that it can be seen in different or different ways (multiple forms) among the people of the population, and the second is that it has a genetic origin and can be transmitted from one generation to another. Microsatellite molecular marker or simple repetitive sequences (SSR)) consist of a short repetitive double sequence (2-6 base pairs) of DNA (Mohammadi et al., 2009). Microsatellite markers are used as common markers for gene localization due to their high polymorphism, coexpressed nature, easy evaluation and high reproducibility. In addition to having advantages such as high frequency and high polymorphism, these markers have a uniform distribution on the genome and their chromosomal location has been identified on genetic maps.