The ZOS7-MYB60 module confers drought-stress tolerance in rice

Shanlan upland rice is an important landrace resource with high drought stress(DS)tolerance.Despite its importance,genes responsible for yield in Shanlan upland rice have yet to be discovered.Our previous study identified a drought-responsive zinc finger protein,ZOS7,as highly expressed in Shanlandao upland rice.However,the function of this gene in controlling drought tolerance remains largely unexplored.In this study,we found that overexpressing ZOS7,a drought-responsive zinc finger protein,in rice increased biomass and yield under drought stress.Co-overexpressing ZOS7 and MYB60,encoding a protein with which ZOS7 interacted,intensified the yield increase.ZOS7 and MYB60 appear to form a module that confers drought tolerance by regulating stomatal density and wax biosynthesis.The ZOS7-MYB60module could be used in molecular breeding for drought tolerance in rice.

Heterogeneous expression of stearoyl-acyl carrier protein desaturase genes SAD1 and SAD2 from Linum usitatissimum enhances seed oleic acid accumulation and seedling cold and drought tolerance in Brassica napus

Flax(Linum usitatissimum L.)is a versatile crop and its seeds are a major source of unsaturated fatty acids.Stearoyl-acyl carrier protein desaturase(SAD)is a dehydrogenase enzyme that plays a key role in oleic acid biosynthesis as well as responses to biotic and abiotic stresses.However,the function of SAD orthologs from L.usitatissimum has not been assessed.Here,we found that two LuSAD genes,LuSAD1 and LuSAD2,are present in the genome of L.usitatissimum cultivar‘Longya 10’.Heterogeneous expression of either LuSAD1 or LuSAD2 in Arabidopsis thaliana resulted in higher contents of total fatty acids and oleic acid in the seeds.Interestingly,ectopic expression of LuSAD2 in A.thaliana caused altered plant architecture.Similarly,the overexpression of either LuSAD1 or LuSAD2 in Brassica napus also resulted in increased contents of total fatty acids and oleic acid in the seeds.Furthermore,we demonstrated that either LuSAD1 or LuSAD2 enhances seedling resistance to cold and drought stresses by improving antioxidant enzyme activity and nonenzymatic antioxidant levels,as well as reducing membrane damage.These findings not only broaden our knowledge of the LuSAD functions in plants,but also offer promising targets for improving the quantity and quality of oil,and the abiotic stress tolerance of oil-producing crops,through molecular manipulation.

Leaf Morphology Genes SRL1 and RENL1 Co-Regulate Cellulose Synthesis and Affect Rice Drought Tolerance

The morphological development of rice(Oryza sativa L.)leaves is closely related to plant architecture,physiological activities,and resistance.However,it is unclear whether there is a co-regulatory relationship between the morphological development of leaves and adaptation to drought environment.In this study,a drought-sensitive,roll-enhanced,and narrow-leaf mutant(renl1)was induced from a semi-rolled leaf mutant(srl1)by ethyl methane sulfonate(EMS),which was obtained from Nipponbare(NPB)through EMS.Map-based cloning and functional validation showed that RENL1 encodes a cellulose synthase,allelic to NRL1/OsCLSD4.The RENL1 mutation resulted in reduced vascular bundles,vesicular cells,cellulose,and hemicellulose contents in cell walls,diminishing the water-holding capacity of leaves.In addition,the root system of the renl1 mutant was poorly developed and its ability to scavenge reactive oxygen species(ROS)was decreased,leading to an increase in ROS after drought stress.Meanwhile,genetic results showed that RENL1 and SRL1 synergistically regulated cell wall components.Our results revealed a theoretical basis for further elucidating the molecular regulation mechanism of cellulose on rice drought tolerance,and provided a new genetic resource for enhancing the synergistic regulation network of plant type and stress resistance,thereby realizing simultaneous improvement of multiple traits in rice.

Overexpression of the peroxidase gene ZmPRX1 increases maize seedling drought tolerance by promoting root development and lignification

Drought is a main abiotic stress factor hindering plant growth,development,and crop productivity.Therefore,it is crucial to understand the mechanisms by which plants cope with drought stress.Here,the function of the maize peroxidase gene ZmPRX1 in drought stress tolerance was investigated by measurement of its expression in response to drought treatment both in a ZmPRX1 overexpression line and a mutant line.The higher root lignin accumulation and seedling survival rate of the overexpression line than that of the wild type or mutant support a role for ZmPRX1 in maize drought tolerance by regulating root development and lignification.Additionally,yeast one-hybrid,Dule luciferase and ChIP-qPCR assays showed that ZmPRX1 is negatively regulated by a nuclear-localized ZmWRKY86 transcription factor.The gene could potentially be used for breeding of drought-tolerant cultivars.

Aβ-Carotene Ketolase Gene NfcrtO from Subaerial Cyanobacteria Confers Drought Tolerance in Rice

Nostoc flagelliforme is a terrestrial cyanobacterium that can resist many types of stressors,including drought,ultraviolet radiation,and extreme temperatures.In this study,we identified the drought tolerance gene NfcrtO,which encodes aβ-carotene ketolase,through screening the transcriptome of N.flagelliforme under water loss stress.Prokaryotic expression of NfcrtO under 0.6 mol/L sorbitol or under 0.3 mol/L NaCl stress significantly increased the growth rate of Escherichia coli.When NfcrtO was heterologously expressed in rice,the seedling height and root length of NfcrtO-overexpressing rice plants were significantly higher than those of the wild type(WT)plants grown on½Murashige and Skoog solid medium with 120 mmol/L mannitol at the seedling stage.Transcriptome analysis revealed that NfcrtO was involved in osmotic stress,antioxidant,and other stress-related pathways.Additionally,the survival rate of the NfcrtO-overexpression lines was significantly higher than that of the WT line under both hydroponic stress(24%PEG and 100 mmol/L H_(2)O_(2))and soil drought treatment at the seedling stage.Physiological traits,including the activity levels of superoxide dismutase,peroxidase,catalase,total antioxidant capacity,and the contents of proline,trehalose,and soluble sugar,were significantly improved in the NfcrtO-overexpression lines relative to those in the WT line under 20%PEG treatment.Furthermore,when water was withheld at the booting stage,the grain yield per plant of NfcrtO-overexpression lines was significantly higher than that of the WT line.Yeast two-hybrid analysis identified interactions between NfcrtO and Dna J protein,E3 ubiquitin-protein ligase,and pyrophosphate-energized vacuolar membrane proton pump.Thus,heterologous expression of NfcrtO in rice could significantly improve the tolerance of rice to osmotic stress,potentially facilitating the development of new rice varieties.

Physiological Response Mechanism and Drought Resistance Evaluation of Passiflora edulis Sims under Drought Stress

In order to explore the response mechanism of Passiflora edulis Sims to drought stress,the changes in morpho-logical and physiological traits of Passiflora edulis Sims under different drought conditions were studied.A total of 7 germplasm resources of Passiflora edulis Sims were selected and tested under drought stress by the pot culture method under 4 treatment levels:75%–80%(Control,CK)of maximumfield water capacity,55%–60%(Light Drought,LD)of maximumfield water capacity,i.e.,mild drought,40%–45%(Moderate Drought,MD)of max-imumfield water capacity,i.e.,moderate drought and 30%–35%(Severe Drought,SD)of maximumfield water capacity,i.e.,severe drought.On the 40th day of drought treatment,13 indices,including seedling growth mor-phology,physiology,and biochemistry,were measured.The results showed that under drought stress,the height and ground diameter of P.edulis Sims gradually decreased with increasing drought stress,and there were signifi-cant differences in seedling height and ground diameter among the treatments.Drought stress significantly inhib-ited the growth of seven P.edulis Sims varieties.The contents of soluble sugar(SS),soluble protein(SP),proline(Pro),and other substances in P.edulis Sims basically increased with increasing drought stress.With the aggrava-tion of drought stress,the malondialdehyde(MDA)content of P.edulis Sims tended to increase to different degrees,the superoxide dismutase(SOD)activity and peroxidase(POD)activity both tended to increase atfirst and then decrease,and the change in catalase(CAT)activity mostly showed a gradual increasing trend.The con-tents of endogenous hormones in P.edulis Sims significantly differed under different degrees of drought stress.With the aggravation of drought stress,the abscisic acid(ABA)content of P.edulis Sims tended to increase,whereas the contents of gibberellin(GA),indoleacetic acid(IAA),and zeatin nucleoside(ZR)exhibited a down-ward trend.A comprehensive evaluation of the drought resistance of seven P.edulis Sims varieties was conducted based on the principal component analysis method,and the results showed that the drought resistance decreased in the order XH-BL>XH-TWZ>TN1>GH1>ZJ-MT>LP-LZ>DH-JW.

Physiological and Transcriptome Analysis Illuminates the Molecular Mechanisms of the Drought Resistance Improved by Alginate Oligosaccharides in Triticum aestivum L.

Alginate oligosaccharides(AOS)enhance drought resistance in wheat(Triticum aestivum L.),but the definite mechanisms remain largely unknown.The physiological and transcriptome responses of wheat seedlings treated with AOS were analyzed under drought stress simulated with polyethylene glycol-6000.The results showed that AOS promoted the growth of wheat seedlings and reduced oxidative damage by improving peroxidase and superoxide dismutase activities under drought stress.A total of 10,064 and 15,208 differentially expressed unigenes(DEGs)obtained from the AOS treatment and control samples at 24 and 72 h after dehydration,respectively,were mainly enriched in the biosynthesis of secondary metabolites(phenylpropanoid biosynthesis,flavonoid biosynthesis),carbohydrate metabolism(starch and sucrose metabolism,carbon fixation in photosynthetic organisms),lipid metabolism(fatty acid elongation,biosynthesis of unsaturated fatty acids,alpha-linolenic acid metabolism,cutin,suberine and wax biosynthesis),and signaling transduction pathways.The up-regulated genes were related to,for example,chlorophyll a-b binding protein,amylosynthease,phosphotransferase,peroxidase,phenylalanine ammonia lyase,flavone synthase,glutathione synthetase.Signaling molecules(including MAPK,plant hormones,H_(2)O_(2) and calcium)and transcription factors(mainly including NAC,MYB,MYB-related,WRKY,bZIP family members)were involved in the AOS-induced wheat drought resistance.The results obtained in this study help underpin the mechanisms of wheat drought resistance improved by AOS,and provides a theoretical basis for the application of AOS as an environmentally sustainable biological method to improve drought resistance in agriculture.

Physiological Analysis of Drought Resistance of Seven Cultivars Spring Wheat in Northern Regions of China

In order to determine the physiological mechanism of drought resistance of northern wheat in China,six drought resistant wheat and one sensitivity to drought wheat were planted in pots.They were subjected to drought treatment and normal water when the plants grew to the 3-leaf stage.Samples were collected at 10,20,30,and 40 days after the drought treatment,respectively.The electrical conductivity,photosynthetic parameters,chlorophyll fluorescence parameters,sugar content,proline content,protein content,and active oxygen scavenging enzyme activity of the plants were detected,and the agronomic traits of the wheat varieties were investigated at maturity.The results indicated that the phenotype and yield-related factors of Darkhan 144 changed little under the drought stress.The relative electrical conductivity of Kefeng 6 and Darkhan 166 was lower under the drought stress,and their cell membrane was less damaged.The Darkhan 144 and Darkhan 166 had higher drought resistance coefficients,and were the wheat varieties with stronger drought resistance.However,the physiological mechanisms of drought resistance of these three wheat were different:Darkhan 144 maintained a higher photosynthetic activity under the drought stress;Darkhan 166 maintained a higher protein content,photosynthetic activity and active oxygen scavenging enzyme activity.In addition,other drought-resistant varieties Kefeng 6,Kefeng 10 and Longmai 26 had a higher content of osmoregulatory substances under the drought stress.

Assessment of molecular markers and marker-assisted selection for drought tolerance in barley(Hordeum vulgare L.)

This review updates the present status of the field of molecular markers and marker-assisted selection(MAS),using the example of drought tolerance in barley.The accuracy of selected quantitative trait loci(QTLs),candidate genes and suggested markers was assessed in the barley genome cv.Morex.Six common strategies are described for molecular marker development,candidate gene identification and verification,and their possible applications in MAS to improve the grain yield and yield components in barley under drought stress.These strategies are based on the following five principles:(1)Molecular markers are designated as genomic‘tags’,and their‘prediction’is strongly dependent on their distance from a candidate gene on genetic or physical maps;(2)plants react differently under favourable and stressful conditions or depending on their stage of development;(3)each candidate gene must be verified by confirming its expression in the relevant conditions,e.g.,drought;(4)the molecular marker identified must be validated for MAS for tolerance to drought stress and improved grain yield;and(5)the small number of molecular markers realized for MAS in breeding,from among the many studies targeting candidate genes,can be explained by the complex nature of drought stress,and multiple stress-responsive genes in each barley genotype that are expressed differentially depending on many other factors.

Evaluation of Inherited Resistance Genes of Bacterial Leaf Blight, Blast and Drought Tolerance in Improved Rice Lines

Improved rice lines were developed frome three parents with the resistance or tolerance to bacterial leaf blight,blast and drought stress,respectively,using single-,double-and three-way crosses.The improved lines were assessed for agro-morphological and yield traits under non-drought stress(NS)and reproductive-stage drought stress(RS)treatments.The mean comparison of traits measured between parent plants and progenies(improved lines)were similar,and there were significant and non-significant differences among the parents and improved lines(genotypes)under NS and RS.Smilarly,there was significant and non-significant differences in the interaction among both parent varieties and improved lines for NS and RS.Cluster and 3D-model of principal component analysis did not generate categorical clusters according to crossing methods,and there were no exclusive crossing method inclined variations under the treatments.The improved lines were high-yielding,disease resistant,and drought-tolerant compared with their parents.All the crossing methods were good for this crop improvement program without preference to any,despite the number of genes introgressed.

Improvement of Upland Rice Variety by Pyramiding Drought Tolerance QTL with Two Major Blast Resistance Genes for Sustainable Rice Production

Varalu is an early maturing rice variety widely grown in the rainfed ecosystem preferred for its grain type and cooking quality.However,the yield of Varalu is substantially low since it is being affected by reproductive drought stress along with the blast disease.The genetic improvement of Varalu was done by introgressing a major yield QTL,qDTY_(12.1),along with two major blast resistance genes i.e.Pi54 and Pi1 through marker-assisted backcross breeding.Both traits were transferred till BC_(2) generation and intercrossing was followed to pyramid the two traits.Stringent foreground selection was carried out using linked markers as well as peak markers(RM28099,RM28130,RM511 and RM28163)for the targeted QTL(qDTY_(12.1)),RM206 for Pi54 and RM224 for Pi1.Extensive background selection was done using genome-wide SSR markers.Six best lines(MSM-36,MSM-49,MSM-53,MSM-57,MSM-60 and MSM-63)having qDTY_(12.1) and two blast resistance genes in homozygous condition with recurrent parent genome of 95.0%-96.5% having minimal linkage drag of about 0.1 to 0.7 Mb were identified.These lines showed yield advantage under drought stress as well as irrigated conditions.MSM-36 showed better performance in the national coordinated trials conducted across India,which indicated that improved lines of Varalu expected to replace Varalu and may have an important role in sustaining rice production.The present study demonstrated the successful marker-assisted pyramiding strategy for introgression of genes/QTLs conferring biotic stress resistance and yield under abiotic stress in rice.

Drought-Tolerant Rice at Molecular Breeding Eras:An Emerging Reality

Rice(Oryza sativa L.)stands as the most significantly influential food crop in the developing world,with its total production and yield stability affected by environmental stress.Drought stress impacts about 45%of the world’s rice area,affecting plants at molecular,biochemical,physiological,and phenotypic levels.The conventional breeding method,predominantly employing single pedigree selection,has been widely utilized in breeding numerous drought-tolerant rice varieties since the Green Revolution.With rapid progress in plant molecular biology,hundreds of drought-tolerant QTLs/genes have been identified and tested in rice crops under both indoor and field conditions.Several genes have been introgressed into elite germplasm to develop commercially accepted drought-tolerant varieties,resulting in the development of several drought-tolerant rice varieties through marker-assisted selection and genetically engineered approaches.This review provides up-to-date information on proof-of-concept genes and breeding methods in the molecular breeding era,offering guidance for rice breeders to develop drought-tolerant rice varieties.

OsMas1,a novel maspardin protein gene,confers tolerance to salt and drought stresses by regulating ABA signaling in rice

Drought and salt stresses,the major environmental abiotic stresses in agriculture worldwide,affect plant growth,crop productivity,and quality.Therefore,developing crops with higher drought and salt tolerance is highly desirable.This study reported the isolation,biological function,and molecular characterization of a novel maspardin gene,OsMas1,from rice.The OsMas1 protein was localized to the cytoplasm.The expression levels of OsMas1 were up-regulated under mannitol,PEG6000,NaCl,and abscisic acid(ABA) treatments in rice.The OsMas1 gene was introduced into the rice cultivar Zhonghua 11(wild type,WT).OsMas1-overexpression(OsMas1-OE) plants exhibited significantly enhanced salt and drought tolerance;in contrast,OsMas1-interference(OsMas1-RNAi) plants exhibited decreased tolerance to salt and drought stresses,compared with WT.OsMas1-OE plants exhibited enhanced hypersensitivity,while OsMas1-RNAi plants showed less sensitivity to exogenous ABA treatment at both germination and post-germination stages.ABA,proline and K+ contents and superoxide dismutase(SOD),catalase(CAT),peroxidase(POD),and photosynthesis activities were significantly increased.In contrast,malonaldehyde(MDA),hydrogen peroxide(H2O2),superoxide anion radical(O2-··),and Na+ contents were significantly decreased in OsMas1-OE plants compared with OsMas1-RNAi and WT plants.Overexpression of OsMas1 up-regulated the genes involved in ABA signaling,proline biosynthesis,reactive oxygen species(ROS)-scavenging system,photosynthesis,and ion transport under salt and drought stresses.Our results indicate that the OsMas1 gene improves salt and drought tolerance in rice,which may serve as a candidate gene for enhancing crop resistance to abiotic stresses.

Increase in root density induced by coronatine improves maize drought resistance in North China

Drought stress caused by insufficient irrigation or precipitation impairs agricultural production worldwide.In this study,a two-year field experiment was conducted to investigate the effect of coronatine(COR),a functional analog of jasmonic acid(JA),on maize drought resistance.The experiment included two water treatments(rainfed and irrigation),four COR concentrations(mock,0μmol L^(-1);A1,0.1μmol L^(-1);A2,1μmol L^(-1);A3,10μmol L^(-1))and two maize genotypes(Fumin 985(FM985),a drought-resistant cultivar and Xianyu 335(XY335),a drought-sensitive cultivar).Spraying 1μmol L^(-1)COR at seedling stage increased surface root density and size,including root dry matter by 12.6%,projected root area by 19.0%,average root density by 51.9%,and thus root bleeding sap by 28.2%under drought conditions.COR application also increased leaf area and SPAD values,a result attributed to improvement of the root system and increases in abscisic acid(ABA),JA,and salicylic acid(SA)contents.The improvement of leaves and roots laid the foundation for increasing plant height and dry matter accumulation.COR application reduced anthesis and silking interval,increasing kernel number per ear.COR treatment at 1μmol L^(-1)increased the yield of XY335 and FM985 by 7.9%and 11.0%,respectively.Correlation and path analysis showed that grain yields were correlated with root dry weight and projected root area,increasing maize drought resistance mainly via leaf area index and dry matter accumulation.Overall,COR increased maize drought resistance mainly by increasing root dry weight and root area,with 1μmol L-^(-1)COR as an optimal concentration.

Drought Tolerance in Mung Bean is Associated with the Genotypic Divergence, Regulation of Proline, Photosynthetic Pigment and Water Relation

Drought is one of the critical conditions for the growth and productivity of many crops including mung bean(Vigna radiata L.Wilczek).Screening of genotypes for variations is one of the suitable strategies for evaluating crop adaptability and global food security.In this context,the study investigated the physiological and biochemical responses of four drought tolerant(BARI Mung-8,BMX-08010-2,BMX-010015,BMX-08009-7),and four drought sensitive(BARI Mung-1,BARI Mung-3,BU Mung-4,BMX-05001)mung bean genotypes under wellwatered(WW)and water deficit(WD)conditions.The WW treatment maintained sufficient soil moisture(22%±0.5%,i.e.,30%deficit of available water)by regularly supplying water.Whereas,the WD treatment was maintained throughout the growing period,and water was applied when the wilting symptom appeared.The drought tolerant(DT)genotypes BARI Mung-8,BMX-08010-2,BMX-010015,BMX-08009-7 showed a high level of proline accumulation(2.52–5.99 mg g^(−1) FW),photosynthetic pigment(total chlorophyll 2.96–3.27 mg g^(−1) FW at flowering stage,and 1.62–2.38 mg g^(−1) FW at pod developing stage),plant water relation attributes including relative water content(RWC)(82%–84%),water retention capacity(WRC)(12–14)as well as lower water saturation deficit(WSD)(19%–23%),and water uptake capacity(WUC)(2.58–2.89)under WD condition,which provided consequently higher relative seed yield.These indicate that the tolerant genotypes gained better physiobiochemical attributes and adaptability in response to drought conditions.Furthermore,the genotype BMX-08010-2 showed superiority in terms of those physio-biochemical traits,susceptibility index(SSI)and stress tolerance index(STI)to other genotypes.Based on the physiological and biochemical responses,the BMX-08010-2 was found to be a suitable genotype for sustaining yield under drought stress,and subsequently,it could be recommended for crop improvement through hybridization programs.In addition,the identified traits can be used as markers to identify tolerant genotypes for drought-prone areas.

PoWRKY71 is involved in Paeonia ostii resistance to drought stress by directly regulating light-harvesting chlorophyll a/b-binding 151 gene

Although the functions of WRKY transcription factors in drought resistance are well known,their regulatory mechanisms in response to drought by stabilising photosynthesis remain unclear.Here,a differentially expressed PoWRKY71 gene that was highly expressed in drought-treated Paeonia ostii leaves was identified through transcriptome analysis.PoWRKY71 positively responded to drought stress with significantly enhanced expression patterns and overexpressing PoWRKY71 in tobacco greatly improved plant tolerance to drought stress,whereas silencing PoWRKY71 in P.ostii resulted in a drought-intolerant phenotype.Furthermore,lower chlorophyll contents,photosynthesis,and inhibited expression of photosynthesis-related light-harvesting chlorophyll a/b-binding 151(CAB151)gene were found in PoWRKY71-silenced P.ostii.Meanwhile,a homologous system indicated that drought treatment increased PoCAB151 promoter activity.Interactive assays revealed that PoWRKY71 directly bound on the W-box element of PoCAB151 promoter and activated its transcription.In addition,PoCAB151 overexpressing plants demonstrated increased drought tolerance,together with significantly higher chlorophyll contents and photosynthesis,whereas these indices were dramatically lower in PoCAB151-silenced P.ostii.The above results indicated that PoWRKY71 activated the expression of PoCAB151,thus stabilising photosynthesis via regulating chloroplast homeostasis and chlorophyll content in P.ostii under drought stress.This study reveals a novel drought-resistancemechanism in plants and provides a feasible strategy for improving plant drought resistance via stabilising photosynthesis.

Genetic dissection of drought resistance for trait improvement in crops

Reliance on agriculture for food security is a constant in all modern societies.Global climate change and population growth have put immense pressure on sustainable agriculture,exacerbating the effects of environmental stresses.Drought is one of the most pressing abiotic stresses that farmers face,presenting an annual threat to crop growth and yield.Crops have evolved extensive morphological,physiological,and molecular mechanisms to combat drought stress.Drought resistance is a polygenic trait,controlled by a complex genetic network and an array of genes working together to ensure plant survival.Many studies have aimed at dissecting the genetic mechanisms underlying drought resistance.Recent studies using linkage and association mapping have made progress in identifying genetic variations that affect drought-resistance traits.These loci may potentially be engineered by genetic transformation and genome editing aimed at developing new,stress-resistant crop cultivars.Here we summarize recent progress in elucidating the genetic basis of crop drought resistance.Molecular-breeding technologies such as marker-assisted selection,genome selection,gene transformation,and genome editing are currently employed to develop drought-resistant germplasm in a variety of crops.Recent advances in basic research and crop biotechnology covered in this review will facilitate delivery of drought-resistant crops with unprecedented efficiency.

Selection of Cowpea [(Vigna unguiculata (L.) WALP] Genotypes for Drought Tolerance Using Selection Indices

Cowpea [(Vigna unguiculata (L.)] is one of the most important arid legumes cultivated for pulse and forage production. However, in cowpea, not much is known about the base index selection method in breeding for drought tolerance. Consequently, the present study has been conducted to: 1) evaluate the yield performance of cowpea genotypes under artificial drought and well-watered condition;2) develop a base index using multiple traits for ranking genotype performance. The experiment was a 25 × 2 factorial laid out in a Randomized Complete Block Design (RCBD) with three replications. The experiment was carried out in the screen house at the Department of Horticulture at KNUST. The result showed that KPR1-96-73, Simbo, CZ06-4-16, Wilibaly and Agyenkwa were high yielding in well-water condition while Ghana Shoba, Sangaraka, NKetewade, Ghana-Shoni and Korobalen were high yielding genotypes in water stress condition. The average yield reduction was 60.6% for grain respectively. The biplot displays revealed four groups among the genotypes tested which was based on their yielding capacity and drought tolerance. In cluster B high yielding and drought tolerant genotypes were identified, high yielding and drought susceptible have been identified in cluster A, low yielding and drought tolerant in cluster D, and lastly low yielding and drought susceptible in cluster C. Genotypes in cluster B, were the best due to the fact that it combines high yield and tolerance to drought. They were Ghana Shoni, Nketewade, Sangaraka and Ghana shoba. These genotypes might be suitably employed in further drought tolerance breeding program of cowpea.

Antibiotic Resistance and Heavy Metals Tolerance in Gram-Negative Bacteria from Diseased American Bullfrog (Rana catesbeiana) Cultured in Malaysia

A total of 140 bacterial isolates have been successfully isolated from various organs of diseased American bullfrog （Rana catesbeiana） cultured in Malaysia. The most frequently isolated bacteria was Edwardsiella spp. （46 isolates） followed by Aeromonas spp. （33 isolates）, Flavobacterium spp. （31 isolates）, and Vibrio spp. （30 isolates）. Majority of the bacterial isolates were found sensitive to furazolidone （85.0%）, chloramphenicol （85.0%）, oxolinic acid （90.0%）, florfenicol （95.0%）, and flumequine （97.5%）. On the other hand, most of the bacterial isolates were resistant to oleandomycin （77.5%） and lincomycin （87.5%）. Nitrofurantoin and flumequine can be inhibited the growth of all of Vibrio spp. whereas all isolates of Edwardsiella spp. were found sensitive to florfenicol and flumequine. Multiple antibiotic resistance （MAR） index were in range of 0.30-0.40, indicating that bacterial isolates from cultured bullfrogs may have received high risk exposure to the tested antibiotics. In addition, 90-100% of the isolates were resistant to copper, cadmium, and chromium. These results provided insight information on tolerance level of bacterial isolates from cultured bullfrogs to 21 antibiotics as well as heavy metals.

Developing transgenic maize(Zea mays L.) with insect resistance and glyphosate tolerance by fusion gene transformation

Using linker peptide LP4/2A for multiple gene transformation is considered to be an effective method to stack or pyramid several traits in plants. Bacillus thuringiensis（Bt） cry gene and epsps（5-enolpyruvylshikimate-3-phosphate synthase） gene are two important genes for culturing pest-resistant and glyphosate-tolerant crops. We used linker peptide LP4/2A to connect the Bt cry1 Ah gene with the 2m G2-epsps gene and combined the wide-used man A gene as a selective marker to construct one coordinated expression vector called p2 EPUHLAGN. The expression vector was transferred into maize by Agrobacterium tumefaciens-mediated transformation, and 60 plants were obtained, 40% of which were positive transformants. Molecular detection demonstrated that the two genes in the fusion vector were expressed simultaneously and spliced correctly in translation processing; meanwhile bioassay detection proved the transgenic maize had preferable pest resistance and glyphosate tolerance. Therefore, linker peptide LP4/2A provided a simple and reliable strategy for producing gene stacking in maize and the result showed that the fusion gene transformation system of LP4/2A was feasible in monocot plants.