Genetics of biochemical attributes regulating morpho-physiology of upland cotton under high temperature conditions

Background Cotton is a strategically important fibre crop for global textile industry.It profoundly impacts several countries’industrial and agricultural sectors.Sustainable cotton production is continuously threatened by the unpre-dictable changes in climate,specifically high temperatures.Breeding heat-tolerant,high-yielding cotton cultivars with wide adaptability to be grown in the regions with rising temperatures is one of the primary objectives of modern cotton breeding programmes.Therefore,the main objective of the current study is to figure out the effective breed-ing approach to imparting heat tolerance as well as the judicious utilization of commercially significant and stress-tolerant attributes in cotton breeding.Initially,the two most notable heat-susceptible(FH-115 and NIAB Kiran)and tolerant(IUB-13 and GH-Mubarak)cotton cultivars were spotted to develop filial and backcross populations to accom-plish the preceding study objectives.The heat tolerant cultivars were screened on the basis of various morphological(seed cotton yield per plant,ginning turnout percentage),physiological(pollen viability,cell membrane thermostabil-ity)and biochemical(peroxidase activity,proline content,hydrogen peroxide content)parameters.Results The results clearly exhibited that heat stress consequently had a detrimental impact on every studied plant trait,as revealed by the ability of crossing and their backcross populations to tolerate high temperatures.However,when considering overall yield,biochemical,and physiological traits,the IUB-13×FH-115 cross went over particularly well at both normal and high temperature conditions.Moreover,overall seed cotton yield per plant exhibited a posi-tive correlation with both pollen viability and antioxidant levels(POD activity and proline content).Conclusions Selection from segregation population and criteria involving pollen viability and antioxidant levels concluded to be an effective strategy for the screening of heat-tolerant cotton germplasms.Therefore,understanding acquired from this study can assist breeders identifying traits that should be prioritized in order to develop climate resilient cotton cultivars.

Comparative analysis of SIMILAR to RCD ONE(SRO)family from tetraploid cotton species and their diploid progenitors depict their significance in cotton growth and development

Background SRO(Similar to RCD1)genes family is largely recognized for their importance in the growth,develop-ment,and in responding to environmental stresses.However,genome-wide identification and functional characteri-zation of SRO genes from cotton species have not been reported so far.Results A total of 36 SRO genes were identified from four cotton species.Phylogenetic analysis divided these genes into three groups with distinct structure.Syntenic and chromosomal distribution analysis indicated uneven distribu-tion of GaSRO,GrSRO,GhSRO,and GbSRO genes on A2,D5 genomes,Gh-At,Gh-Dt,Gb-At,and Gb-Dt subgenomes,respectively.Gene duplication analysis revealed the presence of six duplicated gene pairs among GhSRO genes.In promoter analysis,several elements responsive to the growth,development and hormones were found in GhSRO genes,implying gene induction during cotton growth and development.Several miRNAs responsive to plant growth and abiotic stress were predicted to target 12 GhSRO genes.Organ-specific expression profiling demonstrated the roles of GhSRO genes in one or more tissues.In addition,specific expression pattern of some GhSRO genes dur-ing ovule development depicted their involvement in these developmental processes.Conclusion The data presented in this report laid a foundation for understanding the classification and functions of SRO genes in cotton.

Development of Molecular Marker Linked with Cercospora Leaf Spot (CLS) Disease Resistance in Vigna radiata, Cloning, and Expression for Evaluating Antifungal Activity against Cercospora canescens

We developed a molecular marker for MAS of mungbean resistant varieties against CLS from the consensus sequence(MB-CLsRG)of identified RGAs(MB-ClsRCaG1 and MB-ClsRCaG2).The MB-CLsRG sequence-specific primer pair was used to screen Cercospora leaf spot(CLS)resistant varieties of mungbean in genomic analysis that showed congruency with phenotypic screening.Validation of molecular marker linkage with CLS resistance was performed using rtPCR in transcriptomic analysis.The sequenced PCR products showed 100%homology with MB-CLsRG sequence and putative disease resistance proteins that confirmed the linkage of molecular marker with CLS resistance in mungbean.The antifungal potential of MB-CLsRG gene encoding protein was assessed.The MB-CLsRG gene sequence was cloned in the E.coli expression vector for recombinant protein production.The recombinant protein was then investigated for its in vitro antifungal potential against Cercospora canescens.The in vitro investigation showed strong antifungal activity of recombinant protein as it restricted the growth of fungal mycelial mass.The results validated the linkage of developed marker with CLS-resistant mungbean varieties;therefore,it can be used to screen resistant varieties from a large population in MAS.Moreover,the recombinant protein of the MB-CLsRG gene sequence revealed antifungal potential,which proved the gene sequence could be suitable to use in transgenic plants technology to develop fungal-resistant transgenic crops.

In silico Prediction and Analysis of Potential Off-Targets and Off-Target Mutation Detection in StERF3-Gene Edited Potato Plants

The imperative aspect of the CRISPR/Cas9 system is a short stretch of 20 nucleotides of gRNA that control the overall specificity.Due to the small size,the chance of its multiple occurrences in the genome increases;however,a few mismatches are tolerated by the Cas9 endonuclease activity.An accurate and careful in silico-based off-target prediction while target selection is preferred to address the issue.These predictions are based on a comprehensive set of selectable parameters.Therefore,we investigated the possible off-target prediction and their screening in StERF3 gene-edited potato plants while developing StERF3-loss-of-function mutants using CRISPR/Cas9 approach.The 201 off-targets for the selected targets of the StERF3 gene were predicted,and 79 werefiltered as potential off-targets.Of these 79,twenty-five off-targets showed scores with defined cut-off values<0.5 and were analyzed in Sterf3-edited potato plants compared to wild-type plants.No off-targeting was found to have occurred in edited plants.

Enhancing cotton resilience to challenging climates through genetic modifications

Cotton is one of the most important fiber crops that plays a vital role in the textile industry.Its production has been unstable over the years due to climate change induced biotic stresses such as insects,diseases,and weeds,as well as abiotic stresses including drought,salinity,heat,and cold.Traditional breeding methods have been used to breed climate resilient cotton,but it requires a considerable amount of time to enhance crop tolerance to insect pests and changing climatic conditions.A promising strategy for improving tolerance against these stresses is genetic engineering.This review article discusses the role of genetic engineering in cotton improvement.The essential concepts and techniques include genome editing via clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9(CRISPR-Cas9),overexpression of target genes,downregulation using RNA interference(RNAi),and virus-induced gene silencing(VIGS).Notably,the Agrobacterium-mediated transformation has made significant contributions to using these techniques for obtaining stable transgenic plants.

Genetic effects conferring heat tolerance in upland cotton(Gossypium hirsutum L.)

Background:Climate change and particularly global warming has emerged as an alarming threat to the crop productivity of field crops and exerted drastic effects on the cropping patterns.Production of cotton has been dropped down to one million bales from 1.4 million bales since 2015 in Pakistan due to the increase in temperature at critical growth stages,i.e.,flowering and boll formation.Keeping in view the importance of cotton in the country,this study was conducted to investigate the genetic effects conferring heat tolerance in six populations(P1,P2,F1,F2,BC1 and BC2)developed from cross-1 and cross-2,i.e.,VH-282×FH-142 and DNH-40×VH-259.Results:The results revealed that cross-1 performed better in heat stress as compared with cross-2 for majority of the traits recorded.Boll weight and ginning outturn(GO-T)were highly effected under heat stress and had negative correlation with Relative cell injury(RCI).Boll weight,fiber length,fiber strength and fiber fineness were under the control of non-additive gene action,whereas RCI was controlled by additive gene effects.Lower values of genetic advance coupled with higher values of broad sense heritability for these traits except RCI confirmed the role of non-additive genetic effects.Duplicate types of epistasis were recorded for fiber strength in cross-1 in normal condition.However,complementary type of non-allelic interaction was recorded for fiber strength under normal condition,fiber fineness and RCI under heat stressed condition in cross-1.Likewise,boll weight,GOT and fiber length in populations derived from cross-2 in normal condition were also under the influence of complementary type of non-allelic interaction.Significant differences among values of mid parent and better parent heterosis for boll weight in both normal and heat stress condition provided the opportunity to cotton breeders for utilization of this germplasm for improvement of this trait through exploitation of heterosis breeding.Conclusion:Cross-1 performed better in heat stress and could be utilized for development of heat tolerant cultivar.RCI was under the influence of additive gene action,so one can rely on this trait for screening of large number of accessions of cotton for heat stress.While other traits were predominantly controlled by non-additive gene action and selection based on these should be delayed in later generations.

Polymorphic information and genetic diversity in Brassica speciesrevealed by RAPD markers

Randomly amplified polymorphic DNA(RAPD)is a tremendously convenient approach used to discriminate between Brassica species owing to its accuracy and speed.RAPD primers generate adequate genetic information that can be used in the primer-marker system.In this work,twenty RAPD-PCR based markers were executed to generate polymorphic data,like polymorphic information content(PIC),mean resolving power(MRP),resolving power(RP),effective multiplex ratio(EMR),and marker index(MI)for the first time and genetic distance among and between six Brassica species were calculated.Our results indicated that 20 primers produced a total of 231 scored band and generated 87%polymorphic bands.Average PIC,MRP,RP,MI,and EMR values were 0.088,0.65,6.7,0.78,and 8.9,respectively.PIC showed an overall negative correlation with MRP,RP,MI,and EMR,whereas MRP,RP,and EMR,were positively correlated with each other.Genetic identities ranged from 41.99%(between Brassica napus and Brassica oleracea)to 68.83%(between Brassica campestris and Brassica oleracea).Dendrogram results showed no clustering between species except between Brassica campestris and Brassica nigra.Nevertheless,these results will be helpful to acquire useful information about the markers and their use to determine the genomic structures of Brassica species.Further,based on genetic distance and polymorphic information,new hybrids can be developed for effective oilseed production.

A discussion on cotton transformation during the last decade (2010–2021);an update on present trends and future prospects

The introduction of genetically modified(GM)cotton in 1996 in the US and its worldwide spread later rejuvenated cotton production in many parts of the world.The evolution is continued since then and currently,the 3rd and fourth generation of same GM cotton is grown in many parts of the world.The GM cotton introduced in 1996 was simple Bt cotton that expressed a single Cry1Ac gene,the later generation carried multiple Cry genes along with the genes controlling herbicide tolerance.Current day GM cotton does not only give stable resistance against lepidopteran insects but also facilitates the farmers to spray broad-spectrum herbicides without harming the crop.The evolution of GM cotton is continued both on the basic and applied side and interventions have been introduced during the last decade.Earlier the cotton transformation was limited to Cocker strains which are getting possible in many other varieties,too.It is successful with both gene gun,and Agrobacterium and inplanta transformation has made it a routine activity.Apart from overexpression studies for various purposes including biotic,abiotic,and quality traits,RNAi and genome editing are explored vigorously.Through this review,we have tried to explore and discuss various interventions for improving transformation protocols,the applications of cotton transformation,and future strategies being developed to get maximum benefits from this technology during the last decade.

Improving Growth Performance of Jatropha curcas by Inducing Polyploidy through Colchicine Treatment

This study was planned to assess genetic variation in Jatropha curcas as affected by colchichine treatments. During the experiment, seeds of Jatropha curcas were collected from mature and healthy Jatropha plants and were soaked in different concentrations of colchicines. The second treatment was that of time of soaking (seeds were soaked in different concentrations for different time durations). The results clearly revealed that by increasing the concentration of colchicine treatment, parameters like stomatal density, and size of the guard cells were reduced.

Gene Action Studies in the Inheritance of Economic Traits in Diallel Cross of Lentil (Lens culinaris Medik)

The present study was conducted in 4 × 4 half diallel cross fashion using the genotypes named as NL-2002, NL-2006, Punjab Masoor-2009 and Markaz-2009 for the evaluation of gene action in the inheritance of economic traits of lentil by following the Griffing (1956) Method of analysis. Minirap 13.1 Copyright© 2000, Minitab inc. Method II, Model I was used for the analysis and gene action was computed by variance component analysis using Mixed Model with GLM (general linear model) procedure. The F1 direct crosses along with the parental genotypes were then evaluated by framing the experiment in RCBD in 3 replications. The data were recorded for the characters which are under the study are, plant height (cm), number of primary branches, number of secondary branches, number of pods per plant, number of seed per pod, seed yield per plant, biomass (gram), 100 seeds weight (gram), harvest index (%), yield per plant. The simple ANOVA for all the characters revealed that difference among genotypes was highly significant. As significant differences were present between the genotypes, it permitted subsequent analysis of the data. The Combining Ability (CA) results showed that value of variance for GCA (General Combining Ability) was significantly higher than that of those of the values of variance of SCA (Specific Combining Ability). Hence current results clearly depicted the role of additive type of gene action governing the characters under study.

Genome-wide association analysis for stripe rust resistance in spring wheat(Triticum aestivum L.) germplasm

Stripe rust is a continuous threat to wheat crop all over the world.It causes considerable yield losses in wheat crop every year.Continuous deployment of adult plant resistance(APR)genes in newly developing wheat cultivars is the most judicious strategy to combat this disease.Herein,we dissected the genetics underpinning stripe rust resistance in Pakistani wheat germplasm.An association panel of 94 spring wheat genotypes was phenotyped for two years to score the infestation of stripe rust on each accession and was scanned with 203 polymorphic SSRs.Based on D’measure,linkage disequilibrium(LD)exhibited between loci distant up to 45 c M.Marker-trait associations(MTAs)were determined using mixed linear model(MLM).Total 31 quantitative trait loci(QTLs)were observed on all 21 wheat chromosomes.Twelve QTLs were newly discovered as well as 19 QTLs and 35 previously reported Yr genes were validated in Pakistani wheat germplasm.The major QTLs were QYr.uaf.2 AL and QYr.uaf.3 BS(PVE,11.9%).Dissection of genes from the newly observed QTLs can provide new APR genes to improve genetic resources for APR resistance in wheat crop.

Extraction of DNA suitable for PCR applications from mature leaves of Mangifera indica L.

Good quality deoxyribonucleic acid （DNA） is the pre-requisite for its downstream applications. The presence of high concentrations of polysaccharides, polyphenols, proteins, and other secondary metabolites in mango leaves poses problem in getting good quality DNA fit for polymerase chain reaction （PCR） applications. The problem is exacerbated when DNA is extracted from mature mango leaves. A reliable and modified protocol based on the cetyl trimethylammonium bromide （CTAB） method for DNA extraction from mature mango leaves is described here. High concentrations of inert salt were used to remove polysaccharides; Polyvinylpyrrolidone （PVP） and 13-mercaptoethanol were employed to man＇age phenolic compounds. Extended chloroform-isoamyl alcohol treatment followed by RNase treatment yielded 950-1050 pg of good quality DNA, free of protein and RNA. The problems of DNA degradation,contamination, and low yield due to irreversible binding of phenolic compounds and coprecipitation of polysaccharides with DNA were avoided by this method, The DNA isolated by the modified method showed good PCR amplification using simple sequence repeat （SSR） primers, This modified protocol can also be used to extract DNA from other woody plants having similar problems.