Breeding Strategies for Improvement of Drought Tolerance in Rice: Recent Approaches, and Future Outlooks

Abstract

Rice production is severely limited by drought stress, which causes significant monetary losses. The global climate change is turning into a more significant problem. Enhancing agricultural yield in the drought-prone rainfed areas has become critical in light of the current and projected global food demand. There is a need for rice varieties with drought tolerance in order to achieve the production objective from rainfed areas, and genetic improvement for drought tolerant should be a high priority issue of study in the future. The intricate structure of breeding for drought-tolerant rice varieties makes it a challenging endeavour, and multigenic regulation of drought-tolerant features would be a significant roadblock for the ongoing study. In this chapter, we discussed on the recent crop improvement program for the development of drought-tolerant rice varieties and highlighted the most recent advancements through conventional to molecular breeding level for adaption of cultivars against drought tolerance in rice under different agro-climatic conditions.

1. Introduction

Globally, more than one-third of the world’s population consumes rice e (Oryza sativa L.) as a main staple meal, and a large majority of people, particularly in Asian countries, rely on it for ~80% of their daily caloric needs [1, 2]. In terms of rice production and consumption, Asia is in first place (FAO report, 2020–2021). The tiny root system, thin cuticular wax, and quick stomata closure of rice, however, make it one of the plants most vulnerable to drought [3, 4, 5]. A prerequisite for the effort to achieve self-sufficiency in rice production by 2050 is the creation of high yielding rice varieties with a high level of tolerance and resistance to both biotic and abiotic stressors under adverse climatic conditions [6, 7]. Stress may result from biotic causes like the prevalence of pests, insects, and diseases or abiotic factors such heavy metal toxicity, flooding, salinity, drought, high temperatures, and air pollution, among others. Drought is one of the most destructive abiotic elements [8, 9]. Depending on the plant’s stage of growth, drought stress can result in complete yield losses. Yield losses must be kept to a minimum to aid impoverished rice farmers in emerging nations and ensure food sustainability for the world’s expanding population [9].

The two most significant limiting factors for the low production of rice worldwide are the escalating severity of droughts and the scarcity of high-yielding genotypes that can be grown in drought-prone environments [8]. Due to a lack of suitable rice cultivars and farming methods, rice cultivation is seasonal. Rice cultivation is impacted by the decrease in water supplies brought on by the depletion of important groundwater resources. Due to their immobility, plants have very little chance of escaping the drought state [10]. Severe drought stress can be damaging to plant development at all stages. Low reproductive success for many plant species is caused by the consequences of water deficiency during the reproductive development stage, which can result in male sterility and embryo abortion shortly after pollination [4, 11]. Therefore, understanding how plants respond to the stress becomes vital and primary to designing plants that are resistant to such stress.

Genotype, environment, and the interaction between genotype and environment all have a role in how a plant grows and develops. The biochemical activities that are influenced by environmental influences are also necessary for development [12]. Plants become stressed when environmental conditions are not optimal, which negatively impacts their production, growth, and development. There are two distinct categories of drought conditions: terminal [13] and intermittent [9]. A terminal drought state is brought on by a reduction in the amount of water that is accessible to plants, which causes extreme drought stress and the eventual death of the plant. However, intermittent drought conditions, which happen once or repeatedly during planting seasons, cause plant growth to suffer during the periods of insufficient irrigation. Intermittent drought conditions, in contrast to terminal drought stress, are typically not fatal. Plant survival and ability to retain function during intermittent and terminal drought conditions are key components of drought tolerance or resistance mechanisms [4, 14, 15].

Over the past few decades, study on drought has been increasingly important due to both its rising frequency and its significance for crop output. Nevertheless, it has been difficult to examine drought responses due to the quantitative and complicated character of the drought-tolerant trait [16]. Rice productivity can be increased in a sustainable and economically feasible way by breeding rice cultivars that are tolerant to drought stress [9, 15]. Researchers have tried to breed for drought-tolerant rice plants in the past, but because there aren’t many donors with a high level of drought tolerance, progress is being made slowly. Only a few drought-tolerant variants have yet been identified after screening thousands of samples of germplasm for drought resistance in different parts of the world [17]. The main causes of the limited success are the absence of really drought-tolerant genotypes and the lack of appropriate screening techniques [7, 15]. Nearly 1000 Gene bank accessions originating from 47 different countries were examined for drought tolerance over the course of the previous two decades by researchers at the International Rice Research Institute (IRRI), in the Philippines [2, 18] they have discovered 65 more aus or indica accessions that can withstand drought [19]. In terms of aus accessions, the majority of drought-tolerant varieties are from Bangladesh (19), followed by India (7), while the most of drought-tolerant varieties are from India (16), Bangladesh (3), and Sri Lanka (3) [2]. The use of these rice accessions in next crop improvement projects requires molecular genetics and characterisation for drought tolerance. The most promising sources of genes related to drought that can be employed in the creation of contemporary crop varieties are those cultivars that display great drought resistance [5].

Therefore, one of the most crucial phases in the development of the drought-tolerant crop is knows how plants react to drought stress. The goals of this chapter were (i) to explain how drought stress affects rice plants and to highlight current developments in rice’s physiological, biochemical, and molecular adaptation to drought tolerance (ii) to describe the current process for creating a long-lasting rice variety that is drought-resistant through conventional breeding and the application of biotechnological tools, and (iii) to conduct a thorough analysis of the information that is currently available on drought-resistant genes/QTLs, QTL analysis, gene introgression, and marker-assisted selection.

2. Mechanisms of drought stress and their responses to drought stress in rice

The word “stress” is frequently interpreted physically, as a reaction to various circumstances. Stress is typically an alteration of physiological conditions brought on by elements that seek to compromise the plant’s stability [20]. Low or no precipitation is a climatic characteristic of drought. The majority of the time, drought pressures develops when there is little water in the soil and a constant loss of water through evaporation and transpiration [6]. The term “drought tolerance” refers to a plant’s ability to produce its highest economic yield when water is scarce [21]. It is a complex trait depends on the action and interaction of different morphological, biochemical, and physiological responses are some of the mechanisms that are influenced by genetic variables at various stages are shown in Figure 1 [22]. According to Kumar et al. [22], “drought escape” is defined as the ability of a plant to complete its life cycle before the development of serious soil water deficits. “Drought avoidance” is the ability of plants to maintain relatively high tissue water potential despite a shortage of soil moisture is shown in Figure 1 [23].

Figure 1.

Mechanisms of drought stress and their responses to drought stress in rice. (A) Different responses and mechanism of the rice plants under drought stress; (B) Plant response mechanisms to drought stress.

2.1 Responses of plant morphological traits to drought stress in rice

When rice is subjected to water stress, morphological changes in the early stages of the grain are observed. Normal productivity depends on the timely and ideal establishment of a crop stand. Blighted germination and lowered growth are the primary effects of drought stress [23, 24, 25]. Due to the lack of water, severe reductions in seed germination and growth are seen during drought stress [26]. The germination process is significantly impacted by drought because it prevents water intake and weakens seedlings [26]. Drought stress interferes with water balance, impairs membrane transport, disrupts metabolic processes at the cellular level, and reduces ATP synthesis and respiration, which results in poor seed germination [24]. Water stress causes declines in plant height, leaf area, and biomass, according to a number of reports [25, 27, 28]. Due of the low water potential caused by the drought, leaf growth is inhibited [29]. Crops respond by having poor cell development and reduced leaf area due to disrupted water passage from the xylem towards another cell, as well as lower turgor pressure as a result of water shortage [28]. Under drought-stressed conditions, the anatomy of the leaf and its ultrastructure are altered [30]. These modifications include reduced leaf size, fewer stomata, thick cell walls, cutinisation of the leaf surface, and inadequate conducting system development [21].

Other significant characteristics of plants under drought stress include rolling of the leaves and the beginning of early senescence [31]. According to [25, 28] larger flag leaf area, leaf area index, leaf relative water content, and leaf pigment content have all been used to screen for drought-tolerant varieties of plants. For increasing output while under drought stress, a plant’s root properties are essential. The structure and development of rice root system determine crop function under water stress. By using root mass (dry) and length, it is possible to predict rice output under water stress [32]. On the properties of root growth under water stress, a variety of reactions are seen. A rise in the content of abscisic acid in the roots caused to notice that the length of rice roots increased when under drought stress [33]. Generally speaking, rice cultivars having a deep and voluminous root system are more drought-resistant [34, 35]. For rice, genotypes with a deep root system, coarse roots, the ability to produce numerous branches, and a high root-to-shoot ratio are crucial for drought resistance [35]. Under drought stress, the morpho-physiological traits of rice roots significantly influence shoot growth and total grain output [35]. On the other side, morphological adaptations include enlarged roots with longer root lengths, waxy or thick leaf coatings, fewer epithelial cells, delayed leaf senescence, and more green leaf area [3, 4, 5].

2.2 Responses of plant physiological traits to drought stress in rice

Water stress from a drought or water shortage affects photosynthesis, one of the key metabolic processes that govern crop growth and yield [29]. When there is not enough water available, the stomata shut, lowering the amount of carbon dioxide that reaches the leaves and causing more electrons to be driven into the reactive oxygen species reaction [34, 36]. The decline of photosynthesis is caused by a number of mechanisms, including stomatal closure, turgor pressure loss, reduced leaf gas exchange, and decreased CO2 uptake, which eventually harms the photosynthetic apparatus [8, 29, 36]. Different representations, such as the water potential of the leaf and relative water content (RWC), can be used to show how a plant and water interact [36]. When plants are under water stress, water consumption efficiency is thought to be a crucial factor in determining their ability to produce. It can be viewed as a strategy for enhancing crop production during drought [34]. RWC is a crucial characteristic of water relations in plants and is regarded as the finest integrated measurement of plant water status because it captures changes in the water potential and turgor potential of the plants [8].

In general, the effects of drought stress include a drop in the water content of plants, a reduction in cell length and growth, the closing of stomata, a decrease in gaseous exchange, and the disruption of enzyme-catalysed activities [2]. Additionally, in times of extreme dryness, photosynthesis and metabolism are severely disrupted, which ultimately results in plant death [37]. When compared to cell divisions, drought stress inhibits cell growth [31]. The several biochemical and physiological processes that are impacted by this restriction on plant growth include ion absorption, respiration, photosynthesis, growth promoters, carbohydrate, source-sink relationships, and nutrient metabolism [38]. Chlorophyll content is increased, osmotic potential is decreased, and harvest index is decreased in cells that have been adapted to withstand dryness. Higher stomatal density and conductance, lower transpiration rates, reduced and early asynchrony between female and male flowering and maturation, and improved production, accumulation, assimilation, and yield partitioning are all characteristics of physiological acclimation [6, 17].

2.3 Responses of plant biochemical traits to drought stress in rice

In response to drought stress, plants accumulate organic and inorganic solutes that lower the osmotic potential in an effort to maintain cell turgor. Osmotic adaptations are provided for the plants through the accumulation of osmoprotectants such as proline, glycinebetaine, and soluble sugar [22, 30]. Drought resistance is improved by protein content and profile, as well as a rise in antioxidant activity for scavenging reactive oxygen species [15]. Improved drought response without lowering yield is achieved via tissue- and time-specific expression of drought-responsive features including abscisic acid, brassinosteroids, and ethylene phytohormone pathways etc., [8].

2.3.1 Osmolyte buildup in a drought-stressed environment in rice

The primary process in plants is osmoregulation, and when turgor declines, osmoprotectants accumulate. Under conditions of water scarcity, accumulation of different osmolytes, such as proline, soluble sugar, phenolic, and total free amino acids, increases and plays a significant role in the ability of plants to withstand drought [16, 31]. Plant cells must detect an above- or below-ground incidence of an imbalance between water loss and water availability before their perception may be translated into a cellular stress signal, which is then used to activate drought resistance systems. Plants, which are sessile organisms, have developed a sophisticated signalling system that uses a variety of primary and secondary signal transduction pathways to spread stress messages throughout the entire plant. Since changes in gene expression frequently involve a mix of hormone signals along with the buildup of additional metabolic products including reactive oxygen species, proteins, and other osmolytes, these pathways contain a variety of signalling molecules [16]. Turgor pressure is maintained in dry conditions by the buildup of organic and inorganic solutes, which reduces the osmotic potential in the cytosol. A kind of osmotic adaptation, this metabolic process is highly dependent on the degree of water stress [16, 31]. Proline [39], sucrose [40], glycine betaine [41], and other solutes etc., build up in the cytoplasm as osmotic adaptation happens, encouraging water uptake.

In plants, proline works as an osmolyte in a variety of harmful situations [42]. There are discrepancies between proline accumulation under stress and normal conditions in rice [43]. Comparing dry conditions to well-watered conditions, the proline buildup rises in all rice cultivars [34]. Higher proline buildup is typically connected with greater resistance to drought, and it aids in maintaining leaf turgor and advancing stomatal conductance [22]. Proline content can therefore be used to screen plants for dehydration using biochemical markers [15, 34]. Carbohydrates/soluble sugars are the structural component that provides the energy needed to support plant biomass. Disaccharides, oligosaccharides, and fructans are primarily three forms of water-soluble carbohydrates that play a critical role in stress tolerance under abiotic stress [40]. The balance of numerous physiological activities, including photosynthesis and mitochondrial respiration, depends heavily on soluble sugars [44]. Since plants use a variety of sugar-based coping mechanisms to adapt to environmental stress, sugars play a variety of roles in plants [41]. The availability of mannitol, sorbitol, and trehalose is crucial for the plant’s healthy growth and metabolic operation. Because of the accumulation of soluble sugars that drought causes, the plants are somewhat protected from adverse conditions and even act as osmoprotectants [22, 30, 41].

2.3.2 Antioxidants’ function in drought stress

The plants have an antioxidant defence system that protects them from oxidative harm. Both enzymatic and non-enzymatic antioxidants are present. Antioxidants are essential components of plants that scavenge reactive oxygen species (ROS), and rice that expresses them is more drought-tolerant [2, 5, 44]. The most frequent occurrence when there is a drought stress is an imbalance between the generation and quenching of ROS. In rice, a drought-related imbalance in ROS production and quenching can lead to oxidative damage and negatively impact the life cycle of the plant by reacting with proteins, lipids, and deoxyribonucleic acid [44]. Electron leakage to 1 O2 and the subsequent Mehler reaction that produces ROS have a negative impact on photosynthesis. Due to the negative consequences of the photo-respiratory pathway during drought, excessive levels of superoxide radical (O₂), hydrogen peroxide (H₂O₂), and hydroxyl radical (OH) are formed [44]. These are extremely harmful radicals that cause cellular death by causing lipid peroxidation, protein, and membrane damage to a variety of cell components under drought stress [44]. Therefore, reducing excessive ROS production or increasing antioxidant activity in rice organs is the most effective strategy to improve rice’s ability to withstand drought. Figure 2 illustrates the mode of ROS formation, harmful effects of oxidative stress, cell damage that causes plant death, and several antioxidative systems that scavenge ROS.

Figure 2.

Schematic representation of reactive oxygen species (ROS) damage and antioxidant protection of rice plants under drought stress [44]. APX, ascorbate peroxidase; CAT, catalase; DHAR, Dehydroascorbate reductase; GR, glutathione reductase; GPX, guaiacol peroxidase; MDHAR, monodehydroascorbate reductase; SOD, superoxide dismutase.

The ROS, which comprise hydroxyl free radicals, superoxide radicals, hydrogen peroxide, and singlet oxygen, lead to DNA mutations, lipid peroxidation, protein denaturation, and disturbance of cellular homeostasis. Plants are protected from the harmful effects of ROS by a sophisticated antioxidant system made up of enzymatic antioxidants and non-enzymatic compounds [2]. The enzyme MDHAR [45], DHAR [44], SOD [44], CAT [45], GR [44], APX [44, 46], GPX [25] and ascorbate-glutathione cycle enzyme are examples of enzymatic antioxidants shown in Figure 1. Ascorbate (AsA) [44] and glutathione (GSH) [15, 46] are examples of non-enzymatic antioxidants found in cells shown in Figure 2. Increased drought stress levels cause both enzymatic and non-enzymatic antioxidant activity in rice to rise. A strategy against oxidative stress and an improvement in drought tolerance in rice can be achieved by increasing the expression of the antioxidant system [15, 44]. These antioxidant defence enzymes’ tendency to be more active reveals their protective role in preventing oxidative damage brought on by drought stress [47].

2.3.3 The role of polyamines in drought stress

Rice responds to drought stress by producing small, positively charged molecules called polyamines (PAs) [12, 47]. Plants contain the PAs putrescine (Put), spermidine (Spd), and spermine (Spm). It can interact with several signalling networks and control homeostasis, membrane stabilisation, and osmotic potential and ionic balance. Increased photosynthetic capability, less water loss, and improved osmotic detoxification and adjustment are all directly related to the PA content rise during drought stress [2, 4]. In response to stress, rice produces significantly more putrescine, which encourages the production of spermidine and spermine and eventually protects the plants from dehydration, according to a recent study on the crop [12, 47].

2.3.4 The role of phytohormones under drought stress

Phytohormones are known to play vital roles in regulating various phenomenons in plants to acclimatise to varying drought environment. Abscisic acid (ABA), cytokinins (CK), Jasmonic acid (JA), ethylene (ET), auxins (IAA), gibberellins (GAs) and other major plant hormones are significant in drought response. However, these hormones are usually cross talk with each other to increase the survival of plants in drought condition [2, 23, 48]. Drought stress is experienced as a hydraulic pull brought on by a pressure gradient between the soil and plants as a result of soil drying. The concentration of the signal hormones ABA shifts in response to the perception of a hydraulic force [48, 49]. While other hormones like CKs may be decreased by down-regulating gene expression, degrading via oxidase enzyme activity, or due to stress damage, ABA concentration normally increases in order to communicate the signals associated with drought stress [48, 49]. Since hormone concentration can function independently to confer a signal or it can act in concert with other hormones and/or other signals, these changes are dynamic and complex. Furthermore, the endogenous concentration of a given hormone may be influenced by the duration and severity of drought stress and may differ in different plant organs. One illustration of how hormones interact with one another is the indirect role played by ABA in water stress signalling by suppressing the production of ET [49, 50]. In response to drought, both ABA-dependent and ABA-independent signalling pathways are activated, and a fast buildup of ABA has been linked to improved drought tolerance [2, 4, 49]. In studies of the highly drought tolerant resurrection plants (Craterostigma wilmsii), ABA concentrations were shown to be the most highly affected hormone in response to drought stress [50]. ABA and other hormonal signalling pathways lead to major changes in plant growth, defence responses, and major drought tolerance mechanisms [4].

3. The conventional breeding approaches for improvement of drought tolerance in rice

Normally in conventional breeding methods, grain yield is used as a selection criterion for superior cultivars in drought-prone areas; however this has been demonstrated to be ineffective due to poor heritability and the large impact of genotype by environment interaction [30, 43]. Selection in traditional breeding has steadily moved away from other criteria in favour of physiological qualities since they grow more quickly and depend on genetic variation [43, 51, 52]. However, the main objective of crop breeding is to create high yielding varieties under well water circumstances, and good yielding varieties may continue to produce a high to moderate yield when there is a drought [53]. To put it simply, traditional breeding techniques are crucial for the preservation of germplasm, sexually different parent hybridization, and the emergence of novel genetic characteristics. Using traditional breeding methods, the International Rice Research Institute (IRRI) and the Indian Institute of Rice Research (IIRR), Hyderabad, Telangana, India, have created a wide variety of elite cultivars that are resilient to many diseases and abiotic stresses over the past three decades [2, 53]. In recent years, backcrossing, forced mutation, and pedigree selection have supplanted other traditional breeding techniques as the main ones.

3.1 Pedigree selection

One of the most traditional and popular breeding techniques in rice development is pedigree selection. In particular, if the trait is controlled by important genes, this approach is very suitable for building resistance in rice. The ability to combine numerous genes affecting biotic and abiotic processes is one of the main benefits of pedigree selection [53]. The primary drawback of pedigree selection is that it takes a lot of time and necessitates evaluating numerous lines repeatedly over planting seasons while maintaining a record of the selection criteria. This approach is not appropriate for traits that are influenced by several genes; in this situation, the diallel mating design will be appropriate for selection [53]. Recurrent selection is typically preferred by plant breeders over pedigree selection in the majority of self-pollinating crops, including rice [54]. Figure 3 showed the general selection process for the development of drought-tolerant rice.

Figure 3.

Modified method for conventional yield trail in rice.

3.2 Recurrent selection

In order to increase favourable allele frequencies while preserving genetic diversity, recurrent selection is utilised in varietal improvement. It offers more accurate genetic gains, quicker and more defined breeding cycles, and the creation of extremely diversified breeding lines. This approach, which outperforms the pedigree selection method, has been extensively explored in rice [55].

3.3 Backcross breeding

In order to reduce the genome of the donor parent and consequently increase high recovery of the recipient parent, the backcrossing technique is frequently employed in rice breeding to introduce desirable or target genes controlling a certain trait from donor parent to recipient parent [56]. This method offers a precise and accurate method for creating numerous superior breeding lines. Backcrossing techniques have facilitated the creation of rice cultivars that can withstand drought [56, 57, 58, 59].

3.4 Induced mutation

Since induced mutation has been shown to be effective in the development of improved agronomic traits like an increase in grain yield [57, 58, 59], resistance to pests and diseases, and improvement of physical grain quality, it is used to supplement conventional breeding methods [58]. The creation of gene alleles that are not found in nature is the main benefit of induced mutation, according to [57] who summarised the use of induced mutation with numerous success stories on innovative rice varieties created through induced mutation. Induced mutation is a technique used in plant mutation breeding to create new varieties. Manawthukha rice was exposed to ³⁰⁰ Gy of gamma radiation from a ⁶⁰Co source in Myanmar to test the variety for its ability to withstand drought by withholding irrigation from 90 days after transplant until harvest. Two mutant lines, MK-D-2 and MK-D-3, were determined to be drought resistant after six generations of evaluation and selection by utilising physiological screening procedures [60]. Similar to this, 11 lines with drought-tolerant traits were chosen from an Iranian rice landrace called “Tarom Mahalli” after being exposed to gamma radiation at an optimal dose of ²³⁰ Gy [61]. Induced mutation allowed scientists in Indonesia to create a super green rice mutant that is drought resilient, high producing, and water efficient [62]. Two better lines, MR219-9 and MR219-4, with high production potential and drought tolerance were developed from the common MR219 rice variety in Malaysia [63].

At final, before initiation of new molecular techniques, our understanding of how plants respond to drought at the molecular and whole-plant level has rapidly expanded. There are hundreds of genes that are expressed under drought stress, and some of them have been cloned. The development of drought tolerance often uses a variety of techniques, such as transgenics and gene expression patterns. Proteomics, genome-wide association, stable isotopes, and fluorescence or thermal imaging’s are a few recent techniques that have helped close the genotype–phenotype gap. Rice has developed resilience to drought thanks to genetic engineering and molecular technology, which are the main tools in biotechnological procedures. The most effective and reliable methods to lessen the effects of drought are, in general, the development of genetic resistance.

4. The molecular basis for improvement of drought tolerance in rice

Environmental drought stimuli are detected by yet-to-be-fully-depicted sensors on the membranes, and the signals are then transmitted down through various signal transduction pathways, resulting in the outflow of drought responsive qualities with appropriate gene functions and tolerance to the drought [7, 64]. The phenomenon of drought is complex [22, 65]. So, when it comes to drought tolerance, hybridization and selection techniques cannot provide precise findings. However, the use of DNA markers in molecular investigations can affix the process while still producing precise results. Additionally, by sorting through a large collection of germplasm for drought-tolerant varieties, these molecular markers can be a godsend for further crop improvement. Numerous efforts have been made to identify some qualitative trait loci (QTLs) associated with different attributes [22, 30, 66]. In conclusion, several QTLs for rice drought tolerance have been found Table 1. There have not been many researches on grain yield, though. The vast majority of QTLs in rice have been discovered based on a variety of significant features, such as root and shoot responses, osmotic adjustment, hormonal responses, photosynthesis, and whole plant responses to drought tolerance. DNA studies based on marker-based phenotyping are the main methods used to identify genes involved in rice drought resistance. Despite the advancements, only a few numbers of characteristics have realistically been recognised as having drought resistance capacity [66, 86]. To create transgenic crops with improved resistance to drought stress, it is imperative to identify the candidate genes responsible for plant tolerance under various abiotic stresses [87]. Using genetic engineering (Agrobacterium tumefaciens or gene gun) and hybridization with marker-assisted selection, the gene governing drought tolerance can then be introduced into the genetic background of any suitable cultivar. In this way, molecular breeding can improve crop types and yield varieties, resulting in prolific harvests with high agronomic validity and safety.

4.1 QTLs associated with rice drought tolerance

The plant genome has a number of genes known as QTLs that have extremely precise quantitative properties. Table 2 displays many QTLs connected to various agronomic traits under drought. Earlier molecular genetic studies [52, 66, 97, 98] identified a large number of QTLs linked to various physiological and biochemical traits, but were unable to identify genes that regulate these traits due to poor mapping resolution and weak phenotypic effect [17, 18]. Finding these QTLs associated with selected characteristics aids plant stress screening programmes [99]. Selection of drought-tolerant rice cultivars has made significant use of many QTLs connected to many physiological and growth parameters under drought [52, 96, 99, 100]. Additionally, the classification of QTLs at various stages of rice growth is investigated [52, 96, 99, 100]. Considering yield to be a definite point, continuing research institutes worldwide focus primarily on mapping QTLs for grain yield of rice under drought stress [15]. Thus, specific QTLs for drought tolerance might be found and exploited to create drought-tolerant rice cultivars.

Trait	Pedigree	Marker	Mapping population	No. of QTL	References
Seedling drought resistance	Indica × Azucena	RFLP, AFLP, SSR	RIL	7	[67]
Cellular membrane stability	IR62266 x CT9993	RFLP, AFLP, SSR	DH	9	[68]
Leaf water relations and rolling	Azucena × Bala	RFLP, AFLP, SSR	RIL	13	[69]
Seed fertility, spikelet/panicle, grain yield	Teqing x Lemont	SNP	IL	5	[70]
Root number, thickness, and length	IR58821 × IR52561	AFLP & RFLP	RIL	28	[71]
Root architecture and distribution	IR64 x Azucena	RFLP	DH	39	[72]
Root traits and penetration index	IR1552 × Azucena	SSR	RIL	23	[73]
Deep roots	3 populations	SSR, SNP	RIL	6	[74]
Root penetration, root and tiller number	CO39 × Moroberekan	RFLP	RIL	39	[75]
Root-penetration	Azucena × Bala	RFLP, AFLP	RIL	18	[76]
Grain yield under drought	Two population	SSR	BS	4	[77]
Grain yield in aerobic environments	Three populations	SSR	BS	1	[78]
Yield traits at the reproductive stage	IR64 × Cabacu	SNP	RIL	1	[79]
Yield under stress at reproductive stage	swarna x WAB	SSR	BIL	1	[80]
Heritability for grain yield	CT9993 × IR62266	AFLP	DH	1	[81]
Grain yield under severe lowland drought	R77298 x Sabitri	SSR	BC1	1	[82]
Yield at reproductive stage over environments	Two populations	SSR	BSA	2	[83]
Morphological and physiological traits	IR64 × Azucena	RFLP	DH	15	[84]
Osmotic adjustment and Dehydration tolerance	CO39 × Moroberekan	RFLP	RIL	1	[85]

Table 1.

QTL for yield and yield contributing traits responses under drought stress conditions in rice.

Traits	QTL	Reference
Grain Yield	qDTY1.1, qDTY3.2, qDTY10.1	[77, 88]
	qDTY1.2, qDTY1.3	[89]
	qDTY2.1	[78]
	qDTY2.2, qDTY2.3	[83]
	qDTHI2.3	[34]
	qDTY3.1, qDTY6.1, qDTY6.2	[10]
	qDTY4.1, qDTY9.1, qDTY10.2	[90]
	qDTY9.1A	[10]
	qDTY12.1	[91]
	qPDL1.2, qPNF3.1	[92]
Leaf rolling	qlr8.1	[92]
	qLR9.1	[66]
	qDLR8.1	[10]
Leaf drying	qLD9.1, qLD12.1	[66]
Harvest index	qHI9.1	[66]
	qSf6	[93]
	qPNF3.1	[91]
Spikelet fertility	qSF9.1	[66]
Panicle number	qgy3.1	[94]
Plant height	qPH1.1	[79]
Flowering day	qHGW2.2	[91]
Panicle number, grain weight	qGy7	[95]
Panicle length	qPL-9	[95]
Grain number	qDTY8.1	[77]
Relative water content	qRWC9.1	[66]
Transpiration	qDTR8	[96]
Total dry matter yield	qHGW1	[91]
Days to heading, grain/panicle	qPSS8.1	[79]

Table 2.

QTLs associated with drought tolerance in rice for governing yield and yield contributing traits.

The majority of the QTLs for drought tolerance in rice that have been discovered so far come from non-elite genotypes. The rice plants’ QTL qDTY1.1 is widely applied as a yield characteristic while they are under drought stress [69]. Table 2 lists several significant QTLs that have been discovered in various rice lines, including qDTY2.1 [52], qDTY2.2 [52], qDTHI2.3 [101], qDTY3.1 and qDTY6.1 [96], qDTR8 [100], There are also other SSR markers associated with these QTLs [101]. Therefore, it would be beneficial to use these markers for molecularly screening new rice genotypes for drought tolerance. This would allow for quick and accurate profiling of the rice lines. During the reproductive stage of rice studied the genetic mapping of morpho-physiological traits related to drought tolerance. They reported five QTLs, including qLR9.1, qLD9.1, qHI9.1, qSF9.1, and qRWC9.1, which control, respectively, leaf rolling, leaf drying, harvest index, spikelet fertility, and relative water content in rice [66].

4.2 Rice drought tolerance via transgenic/genetic engineering and genetic methods

The production of several protein classes, such as transcriptional factors, molecular chaperones, enzymes, and other functional proteins, by plants has allowed them to create dependable routes or signalling chains for stress [101]. These proteins increase the ability of plants to withstand or fight drought. In reality, these genes (regulatory elements and proteins) have been discovered using various genomic techniques in numbers of hundreds or even thousands. As shown in Table 3, these genes have been integrated into the rice genome to investigate their impact on drought improvement by overexpression or suppression. In rice, many transcription factors that are encoded by WRKY genes regulate various biological processes. In plants, zinc finger proteins are widely distributed, especially those that control stress responses. Both monocotyledons and dicotyledons have the WRKY genes, which are widely dispersed in plants. Numerous WRKY genes have regulatory functions that can be positive or negative in how plants react to various abiotic stressors [4, 116].

Gene	Function	Reference
DRO1	Induces root elongation and deeper rooting	[102]
OsDREB1F	Maintains ABA-dependent signalling pathway	[103]
OsDREB2B	Root length and number of root increment	[103]
CYP735A	Maintains cytokinin level	[104]
OsNAC5	Enhances root diameter and grain yield	[105]
SNAC1	Enhances spikelet fertility	[16]
OsbZIP23	Increases grain yield	[106]
AP37	Enhances seed filling and grain weight	[107]
OsbZIP46	Increases grain yield	[108]
OsbZIP71	Enhances seed setting	[109]
EcNAC67	Increases relative water content, delays leaf rolling, higher root and shoot mass	[110]
DsM1	Helps in reactive oxygen species scavenging, maintains drought tolerance at the seedling stage	[111]
OsPYL/RCAR5	Induces stomatal closure, regulates leaf fresh weight	[35]
OsWRKY47	Relatively low yield reduction	[112]
AtDREB1A	Osmolyte accumulation, chlorophyll maintenance, higher relative water content and reduced ion leakage	[111]
TlOsm	Maintains growth, retains higher water content and membrane integrity and improves survival rate	[113]
OsMIOX	Higher reactive oxygen species scavenging enzyme activity and proline content	[114]
Coda	Better yield, higher photosystem II activity, increased detoxification of reactive oxygen species	[115]
OsTPS1	Higher trehalose and proline accumulation	[49]
OsCPK9	Increases drought tolerance through enhanced stomatal closure and better osmoregulation in transgenics	[111]
OsNAC10	Increases tolerance to drought at vegetative stage, enlarges roots and improves grain yield	[105]

Table 3.

Genes associated with different mechanisms of drought tolerance in rice.

By decreasing stomata density and enhancing stomata closure, rice zinc-finger protein (dst mutant) demonstrated increased drought and salt tolerance. However, DST non-mutants alter H2O2 homeostasis, which has an adverse effect on stomata closure [116]. By improving drought tolerance, overexpression of the zinc finger protein OsZFP252 demonstrated 74–79% greater chances of survival. Additionally, proline and soluble sugar buildup are increased [127].

About 5000 genes are up regulated and 6000 genes are down regulated in rice after drought stress exposure [18, 128]. Table 4 lists a few of the genes and their associated roles in rice drought tolerance. These genes are divided into three main categories, including those related to membrane transport, signalling, and transcriptional regulation [30, 35]. Under drought stress, they regulate the majority of rice’s biochemical, physiological, and molecular systems [8, 64] According to research by [22, 30] numerous genes and transcription factors exhibit differential expression in rice and are employed for transgenic plants under drought stress. The majority of the genes that are controlled by drought have both ABA-dependent and ABA independent regulation mechanisms [8, 64]. OsJAZ1 reduces drought tolerance in rice by ABA signalling, which coordinates the plant’s responses to drought stress-related growth and development [129]. Additionally, some genes are linked to osmoregulation and late embryogenesis abundant (LEA) proteins, which confer water shortage tolerance in rice [30, 64]. Other genes include OsPYL/RCAR5 and EcNAC67 delay leaf rolling and cause increased root and shoot mass in rice under situations of water deficiency.

Gene action	Gene	Promoter	Gene transfer methods	Phenotype	References
Genes encoding enzymes that synthesise osmotic and other protectants
Polyamine synthesis	ADC	Ubi-1	Biolistic	Improved drought tolerance by producing higher levels of putrescine and spermine synthesis	[12]
abscisic acid Metabolism	CaMV35SP	DSM2	Agrobacterium	Oxidative and drought stress resistance and increase of the xanthophylls and non-photochemical quenching	[117]
Amino acid metabolism	OsOAT	Ubi1	Agrobacterium	Improve drought tolerance and increase seed setting	[118]
Reactive oxygen species	OsSRO1c	Ubi1	Agrobacterium	Oxidative stress tolerance and stomata closure regulation	[118]
Protoporphyrinogen oxidase	PPO		Agrobacterium	Less oxidative damage, and drought tolerance	[119]
Trehalose synthesis	OsTPS1	Actin1	Agrobacterium	Tolerance of rice seedling to drought, cold, and high salinity	[120]
Late embryogenesis abundant (LEA) related genes
LEA protein gene	HVA1	Actin1	Agrobacterium	Cell membrane stability, higher leaf relative water content and increase in growth under drought stress.	[121]
	HVA1	Actin1	Agrobacterium	Drought and salinity tolerance	[122]
	OsLEA3-2	CaMV35S	Agrobacterium	Drought resistance and increase grain per panicle	[117]
Various regulatory genes
Transcription factor	AP37	OsCc1	Agrobacterium	Improve growth performance under drought stress	[107]
	OsbZIP23	Ubi1	Agrobacterium	Wide spectrum to salt, drought tolerance and yield improvement	[113]
	OsbZIP72	CaMV35S	Agrobacterium	Drought resistance and ABA sensitivity	[123]
Harpin protein	Hrf1	CaMV 35S	Agrobacterium	Drought resistance through ABA signalling and antioxidants, and stomata closure regulation	[124]
Jasmonate and ethylene-responsive factor 1	JERF1	CaMV35S	Agrobacterium	Drought resistance	[124]
Ethylene-responsive factor 1	TSRF1	CaMV35S	Agrobacterium	Enhances the osmotic and drought tolerance	[125]
Stress/zinc finger protein	OsiSAP8	CaMV35S	Agrobacterium	Tolerance to salt, drought and cold stress	[126]

Table 4.

Drought tolerant gene that has been tested in rice transferred through genetic engineering/transgenic methods.

The gene DRO1 stimulates root elongation and deeper rooting in transgenic rice [35, 102, 110]. The overexpression of OsDREB2B, CYP735A, and OsDREB1F in rice under drought stress also increases root morphological adaptations [35]. Rice has a DREB2-like gene called OsDRAP1 that confers drought tolerance, according to [116]. It is essential to increase grain yield in rice during droughts, and transgenic methods are used to do this by introducing genes such OsNAC5, OsLEA3--1 [130], OsbZIP71 [109], OsWRKY47 [112], OsbZIP46 [131]. By examining genes like EDT1/HDG11, AtDREB1A, OsMIOX, and OsTPS1, as well as osmolytes accumulation, greater antioxidant enzyme activity, and enhanced photosynthesis, it has been found that transgenic rice has improved water use efficiency [120]. Through improved osmoregulation and stomatal closure, OsCPK9 increases transgenic plants’ ability to withstand drought [132]. Under extreme drought and salinity conditions, transgenic plants’ survival is improved by overexpressing OsDREB2A. [133]. CDPK7 and CIPK03/CIPK12 regulate a number of regulatory proteins, signal transduction pathways, and protein kinases in rice [106]. Under drought stress, OsITPK2 carries decreased levels of inositol triphosphate and ROS homeostasis in rice [117].

The WRKY genes respond to drought stress and are crucial for plant development [2]. Under laboratory or glass house circumstances, various genes have been tested for their ability to confer drought resistance in rice using transgenic techniques. Prior to being used in molecular breeding programmes, these genes should be tested in the field. Trehalose, often referred to as tremalose or mycose, is a key component of abiotic stress, including cold and drought. It protects against stress, stabilises proteins against denaturation, and also stores carbs. Trehalose-6-phosphate synthase (TPS) and trehalose-6-phosphate phosphatase (TPP) are the two primary enzymes that catalyse the manufacture of trehalose in plants; buildup of trehalose in rice has been shown to increase drought tolerance. An increase in trehalose, improved drought tolerance, and decreased photo oxidation in the rice plant under cold and salt stress were seen when a fusion TPP/TPS gene from Escherichia coli (otsA and otsB) was introduced into rice [134]. In conclusion, this study indicated that engineering drought-tolerant genes into rice’s genetic background is promising, provided that a drought-inducible promoter is employed to achieve successful outcomes.

4.3 Marker-assisted selection (MAS) for rice drought tolerance

To find novel genotypes with desirable drought-tolerant features and related genes/loci, the natural genotypic variation in rice can be studied [135, 136]. Through MAS, these novel genotypes can be used in conventional breeding programmes to create rice varieties that are drought tolerant. Breeding programmes are designed to create high yield lines with enhanced quality metrics and then to introduce the cultivars for agricultural use. Drought tolerant rice genotype breeding has been studied in the past [17, 22, 52, 66, 77], but the success rate has been far below expectations due to the challenge of finding suitable donors with a higher tolerant level as well as the nature of its environment-specific nature.

MAS offers the most accurate, environmentally-friendly, fast and economical method of developing superior rice varieties with a certain degree of resistance or tolerance to drought. The IRRI has been the main site for the majority of the marker-assisted breeding techniques used to create drought-tolerant rice varieties in the past 10 years [22, 137]. Several QTLs for drought tolerance in rice are introduced into top cultivars utilising marker-assisted breeding techniques [17]. In the high-yielding variety IR64, they have successfully incorporated QTLs including qDTY9.1, qDTY2.2, qDTY10.1, and qDTY4.1 using a marker-assisted backcrossing method [17]. With the pyramiding of three QTLs, qDTY2.2, qDTY3.1, and qDTY12.1, [138] created the elite Malaysian rice cultivar MR219 that is drought-tolerant. Three QTLs were incorporated into the development of the rice variety TDK1 by [52] for high yield during drought (qDTY3.1, qDTY6.1 and qDTY6.2). Only as a limitation has drought become more significant, and so far no practical steps have been taken to create rice types that are drought tolerant. A large number of high-yielding cultivars, including Swarna, Samba mahsuri, and IR36, which were previously suggested for cultivation in irrigated regions, have been adopted in the drought breeding effort. Because the aforementioned high yielding varieties cannot withstand repeated droughts, significant loss in rice production is observed when these varieties are cultivated by farmers in rainfed ecosystems during the recurrent drought phase [3]. Therefore, improved special rice varieties with high yields during drought and adaptation to a wide range of unfavourable climatic circumstances in the future require additional focus.

5. Future outlooks

The process of developing drought tolerance in rice is challenging and necessitates a thorough understanding of the different morphological, biochemical, physiological, and molecular characteristics. Despite the impressive advancements made by marker-assisted breeding, there are still a number of major obstacles to molecularly breeding rice for drought tolerance. Additionally, the multigenic regulation and complex nature of drought-tolerant characteristics would be a significant roadblock for ongoing and upcoming research in the field. The complex phenomena of rice crop maintenance during drought conditions are governed by the interactions of a number of factors. Transgenic methods are essential for enhancing rice’s agronomic qualities and production characteristics, and they would effectively advance the breeding programme for drought resistance. It is important to understand how these genes react in the presence of drought in the field because several genes have been investigated for their ability to confer drought tolerance in rice under laboratory conditions. Even while substantial basic research is being conducted, we still know relatively little about the mechanism behind the whole-plant stress response. Therefore, we must look into how differentiated cells, tissues, and organs respond to stress and make meaningful connections between the data. In order to improve our understanding of drought tolerance and to promote the genetic improvement of drought-tolerant rice varieties, crop breeding can employ advancements in new technologies related to crop physiology, molecular genetics, and breeding methodologies in an integrated manner.

6. Conclusion

Evaluation for drought resistance is made more difficult by the dynamic and highly variable timing, persistence, and intensity of drought stress under natural settings. Due to their connections to drought stress, abiotic stressors including salt and high temperatures should also be evaluated in conjunction with drought resistance. Many attempts have been made using the numerous QTLs for drought resistance that have been found in rice. High throughput genotyping is now achievable thanks to recent developments in functional genomics, which aid in the identification of key QTL linked to drought tolerance. Therefore, a better knowledge of the genetic basis of drought resistance will be made possible by the successful cloning of these QTL for drought features. The most practical use of drought resistance QTLs, however, is to execute marker-assisted selection based on pyramiding of advantageous QTL alleles to generate drought-resistance in rice utilising recently developing breeding approaches like GWS and MARS. Many genes have been discovered and exploited for enhancing drought resistance via transgenic methods, although the majority of the study was done in glasshouses. The genes that have been shown to be drought tolerance should therefore undergo additional field testing due to the complexity of field settings before being included in the breeding programme. Similar to this, most studies on drought resilience concentrate on the above-ground features, leaving a significant gap for below-ground traits, mostly because phenotyping is challenging. Due to their significant functions in regulating growth and stomata under drought circumstances, root flexibility and architecture should receive enough consideration in studies of drought tolerance.

Acknowledgments

All authors offered equal suggestions on various drafts of the manuscript preparations.

Conflict of interest

The authors declare no conflict of interest.