The new breeding lines are then tested in different soil types in different climatic zones within the regions of release, to ensure no yield penalty of the salt-tolerance gene.This approach of crossing and selection is usually done using molecular markers: DNA fragments that are associated with the trait.Selection for the trait itself is more laborious and expensive.Conventional breeding—For centuries, farmers in countries with extensive soil salinity have long been selecting best yielding crops for their land, as have the more recent commercial breeding companies.If their soil contains salt, they have selected salt-tolerant material without specifically intending to do so.An example is the salt-tolerant bread wheat Kharchia, which forms the basis of most of the salt-tolerant bread wheat germplasm released in India and Pakistan.Kharchia 65 is a land race developed from selections in farmers’ fields in the sodic-saline soils of the Kharchi-Pali area of Rajasthan.We do not yet know the physiological or molecular basis of the salt tolerance of Kharchia.For bread wheat a summary by Naeem et al.listed 14 varieties or land races under commercial production in India, Pakistan, Egypt and China.All of these were produced by conventional breeding.For rice, derivatives of the land races Pokkali or Nona Bokra which occur in the coastal regions of southern India have formed the basis of salt-tolerant rice cultivars.Ismail and Horielist 27 cultivars that have been released for salt tolerance between 2007 and 2014 for Bangladesh, the Philippines and India.These have been developed by conventional selection and breeding.The two most significant cultivars are CSR 36 for salt-affected soils in India, and BRRI Dhan 10 for soils inundated by seawater in coastal Bangladesh.We know the molecular basis of some of this salt tolerance: the presence of specific alleles of the Na+ transporter OsHKT1;5 that enhance Na+ exclusion.

These were identified in Nona Bokra as the QTL SKC1 and identified in Pokkali as the genomic region Saltol which encompasses OsHKT1;5.Molecular markers are now being used to accelerate breeding and to pyramid salt tolerance with other traits relevant to saline soils such as water logging tolerance.Trait-based breeding—A lack of fast and reliable screening methods has been the major limitation to exploring large germplasm collections,round plastic pots selecting genotypes with greater salt tolerance than the current cultivars, and introducing the salt tolerance into breeders’ advanced breeding lines for release of a new salt-tolerant cultivar.Munns and James summarized the various methods used in the laboratory or glasshouse to select for salt tolerance, along with their advantages and disadvantages.The simplest method is that of screening at germination as it is such a quick and easy test for large numbers of genotypes.However, for most species there is little or no correlation between genotypic differences in germination and genotypic differences in later growth or yield.The most reliable and useful method has been to measure rates of Na+ or Cl
accumulation in leaves, selecting individuals with low rates of accumulation.Ideally, biomass or grain yield should be the ultimate criterion for salt tolerance.Selections of various genotypes of pasture species like clover or alfalfa can conveniently be done in hydroponics or sand cultures with added salt, as cuts can be made every 6–8 weeks for replications.Cereals are more difficult to assess as grain yield needs to be measured in saline soil in the field, as does the yield of perennial horticultural species like citrus and grapevine.However, field experiments are plagued by heterogeneities in soil texture and surface elevation and its associated effect on soil salinity and compaction over short distances by influencing soil water deficits or water logging.This heterogeneity makes validation of breeding trials difficult as soil salinity varies greatly over area and depth.Soil salinity under each of a thousand or so breeding plots needs to be measured by electromagnetic induction with a simple-to-use meter such as Geonics EM38 after calibration.Incorporation of plot EC as a co-variant in the statistical analysis was essential to finding durum wheat genotypes and bread wheat and barley genotypes with higher yield in saline soil.

Over the last 20 years, selection of new salt-tolerant germplasm and its use in subsequent breeding has depended on traits and molecular markers for traits, which can be obtained from genetic analysis as Quantitative Trait Loci or by Genome Wide Association Studies.For many crop species, genetic variation in ion exclusion correlates highly with salt tolerance, and screening based on the measurement of ion accumulation in leaves is the most precise and effective form of selection, being quantitative and non-destructive.Examples include Na+ exclusion from leaves of durum wheat and rice.As an example, we describe a successful project on introduction of genes for salt tolerance from a wheat relative into a durum wheat cultivar, using molecular markers for the trait of Na+ exclusion.Durum wheat lacks the gene for Na+ exclusion found in bread wheat.Using the screening method of Na+ exclusion from leaves among 60 durum wheat relatives, Na+ exclusion equal to bread wheat was found in an unusual durum genotype named Line 149.Line 149 was crossed with the durum cultivar Tamaroi which had five times the leaf Na+ concentration and subsequent genetic analysis showed that Na+ exclusion was due to two genes that were named Nax1 and Nax2.Further crossing enabled separation of the two genes, which were identified as HKT1 transporters.Field trials in multiple sites showed that Nax2 increased yield on highly saline soil by 25% without affecting yield on better soils.However, Nax1 had a yield penalty that outweighed its advantage as a Na+ excluder.This yield penalty had not been obvious in glasshouse trials but became significant in the field.Phenomics—For crop species where a trait is multi-genic and covering different chromosome regions, molecular markers have limited value and selection is driven by phenomics.High-throughput phenotyping methods, now employed in the field as well as in the laboratory, allow large numbers of plants to be screened efficiently with limited handling and labor.Screening for salt tolerance in species which do not have a selectable salt-specific trait is only feasible using non-destructive methods.Such methods include biomass growth as assessed by photosynthesis, stomatal conductance, chlorophyll fluorescence and spectral reflectance.

Using color imaging along with nondestructive measurements of the leaf area and growth rate of each plant, it is possible to separate the effects of salinity on new leaf production from the acceleration of senescence and death of old leaves.Imaging allows the short-term osmotic effects on plant growth to be distinguished from the longer-term ionic effects.Infrared thermography is a widely used phenomic tool to detect differences between genotypes in soilless culture, pots, and field plots.In addition, hyperspectral imaging is used to quantify differences in water status and photosynthetic capacity and to detect genotypic differences in salinity tolerance, for example,hydroponic bucket among wheat cultivars after anthesis.Most candidate genes so far discovered and proven to be part of the mechanism of salt tolerance are membrane transporters for Na+, K+ or Cl.Few transcription factors have a known function, either in the downstream target genes, or the cells or tissues in which they operate.Genes involved in signaling pathways are not known to be specific for salinity but have commonalities with other abiotic stresses that reduce growth rate like drought, heat and cold.Transgenics—Use of the Arabidopsis genome has greatly accelerated the sequencing and functional analysis of candidate genes.In total there have been about 7300 papers on salt tolerance involving Arabidopsis.For the six main crop plants there are 9200.How much of this work has led to improving salt tolerance of crops in the field? A summary of 27 genes that have been over expressed in various crop species with “reported plant transgenic performance during salt stress” is listed by Roy et al.in their table 1, but with three exceptions, these transgenics have not been tested in the field or handed over to commercial plant breeders.In a review of genetic engineering for salinity tolerance in wheat , a list of 45 publications on wheat transformed with genes from other species, or other species transformed with genes from wheat, showed only one that included performance in the field; over expression of AtNHX1 improved grain yield of bread wheat.A notable success story is with barley: over expression of AVP1 increased biomass and yield in both non-saline and saline soil.Over expression of genes for accumulation of organic molecules that act as osmolytes such as proline have been studied for decades, but no cultivar has been released with enhanced proline accumulation that improves yield on saline soils.To date, QTL continues to be the main tool of genetic analysis for breeders, yet very few pre-breeding efforts have led to production of salt-tolerant cultivars.Similarly, the early optimism for GWAS to discover new loci for salinity tolerance and their subsequent utilization in varietal development is still not realized.

Success in has been hampered by lack of quantitative and repeatable measurements of the value of the trait to plant growth and yield in saline soil and selection of the best parents for QTL analysis or genotype array.Further research into selection techniques and germplasm diversity is needed.Key genes for Na+ transporters presented in Section 10.2 should be studied using species other than Arabidopsis.Crop species that are amenable to transformation and do not have complex genomes should be used.Omics methodologies should use relevant treatments, such as a gradual and moderate salt stress, not a severe and sudden one.Osmotic shocks cause plasmolysis and induce the synthesis of enzymes that repair the trauma caused to cells by their sudden shrinkage which may take at least 24 h to repair.Gene expression patterns are very different when the stress is imposed gradually compared to a salt shock.Cell-specific and tissue-specific expression is critical for the function of transporters and transcription factors, so studies should consider this should, for example, separately analyze growing from mature tissues.As take-up of genes for salt tolerance by commercial crop breeders has been so slow, and few studies arising with model plants such as Arabidopsis have been validated in the field, there is a high priority to engage plant breeders at an early stage of the project, working along with physiologists, molecular biologists and agronomists.Only then will molecular biology translate to the field and reach crop production targets.There are clear opportunities to make substantial yield gains by targeting basic strategic research, especially by utilizing pre-breeding results of undomesticated varieties, to improve abiotic stress tolerance of crops.Additional recommendations for future research include to use pre-breeding approaches seeking salt tolerance traits, rather than focus on model plants such as Arabidopsis.Also, while research at the cell level is likely to advance our physiological understanding of salt tolerance mechanisms, in parallel significant investments should be made at the field-level, employing the latest in phenotyping methodology.Summary: Unexplored and under-utilized biodiversity exists within crop species and their close relatives, which could be used to improve germplasm for crop production on salt-affected land, without resorting to GM methods that are at present unaccepted in many countries.Ongoing advances in rapid generation turnover, improved phenotyping, envirotyping and analytical methods can increase the rate of genetic gain in breeding.Further understanding of mechanisms at the molecular and physiological level will complement these new technologies and provide farmers with alternatives to increasing crop production on saline land.While genetic improvements cannot provide a permanent solution to increasing soil salinity, and salt-tolerant crops cannot de-salinize the land, a 10% increase in yield may double the famer’s profits, where the profit margin is small.In most of the salt-affected regions with dominance of sodium salts, salinity and sodicity are related, but they are different in terms of their effects on soil environments.“Salinity,” usually measured as total soluble salt concentration, affects plant growth and productivity through osmotic effects and ion toxicity or deficient effects on plant physiological processes.“Sodicity,” generally defined by soil ESPor SARof soil solution, causes constraints to plant growth through its effects on soil physical properties.Natural climatic and soil processes can lead to the formation of sodic soils from saline soils.In irrigated agriculture, the use of sodium containing waters leads to sodic soils by the adsorption of sodium by soils.Sodic soils with low salt concentration undergo structural degradation when wet because of swelling and clay dispersion, causing reduced water and air transport in near-surface soils and to limitations in soil aeration and infiltration.The effects of sodicity on soil physical properties are modified by soil salinity levels.