The Characterization of G-Quadruplexes in Tobacco Genome and Their Function under Abiotic Stress.

Tobacco is an ideal model plant in scientific research. G-quadruplex is a guanine-rich DNA structure, which regulates transcription and translation. In this study, the prevalence and potential function of G-quadruplexes in tobacco were systematically analyzed. In tobacco genomes, there were 2,924,271,002 G-quadruplexes in the nuclear genome, 430,597 in the mitochondrial genome, and 155,943 in the chloroplast genome. The density of the G-quadruplex in the organelle genome was higher than that in the nuclear genome. G-quadruplexes were abundant in the transcription regulatory region of the genome, and a difference in G-quadruplex density in two DNA strands was also observed. The promoter of 60.4% genes contained at least one G-quadruplex. Compared with up-regulated differentially expressed genes (DEGs), the G-quadruplex density in down-regulated DEGs was generally higher under drought stress and salt stress. The G-quadruplex formed by simple sequence repeat (SSR) and its flanking sequence in the promoter region of the NtBBX (Nitab4.5_0002943g0010) gene might enhance the drought tolerance of tobacco. This study lays a solid foundation for further research on G-quadruplex function in tobacco and other plants.

[Genetic background of lily germplasm resources based on SSR markers].

To study the genetic background of lily (Lilium spp.) germplasm resources, and accurately evaluate and select excellent germplasm for genetic improvement of lily, we analyzed the genetic background of 62 lily germplasm accessions from 11 provinces of China by using simple sequence repeat (SSR) molecular markers. The results showed that 15 out of 83 pairs of lily SSR primers were polymorphic. A total of 157 allelic loci were amplified, with the number of alleles per locus ranging from 5 to 19 and the average number of effective alleles per locus being 4.162 8. The average observed heterozygosity and expected heterozygosity were 0.228 2 and 0.694 1, respectively. The average polymorphic information content was 0.678 8. The average Nei's diversity index and Shannon's information index were 0.694 1 and 1.594 9, respectively, indicating that the tested lily germplasm had high genetic diversity. The 62 germplasm accessions were classified into 5 groups by the unweighted pair group method with arithmetic mean (UPGMA) and into 3 groups by the principal component analysis. The two analyses revealed a geographic correlation among different groups. The majority of lily germplasm accessions from the same source tended to cluster together. The population structure analysis classified the lily accessions into 4 populations and 1 mixed population. The above results provide a theoretical basis and genetic resources for the precise identification and breeding of lily germplasm resources.

Genetic Diversity and Reproduction Trends of Phytophthora infestans in Estonia: EU_41_A2 Detected without an Indication of Clonal Reproduction.

This study explores the population dynamics of Phytophthora infestans in Estonia from 2005 to 2022, focusing on genetic diversity and potential shifts in reproductive strategies. In total, 153 P. infestans isolates were collected throughout Estonia over ten growing seasons. Genotyping revealed considerable genetic diversity, with most isolates not corresponding to known multilocus genotypes (MLGs). Still, instances of invasive clonal lineages were observed, notably EU_41_A2. The data indicate the likelihood of random mating rather than clonal reproduction in all the analyzed years. The principal coordinate analysis (PCoA) results revealed no distinct clustering among the sampling years. Statistical analysis and the minimum spanning network (MSN) indicated low genetic differentiation between years with minimal fluctuations in allele frequencies. The continuous monitoring of P. infestans populations is essential for detecting any changes from the current evolutionary trajectory and implement effective disease management strategies, especially considering the recent emergence of EU_41_A2 in the Nordics and the potential impacts of climate change.

The chickpea WIP2 gene underlying a major QTL contributes to lateral root development.

Lateral roots are a major component of root system architecture, and lateral root count (LRC) positively contributes to yield under drought in chickpea. To understand the genetic regulation of LRC, a biparental mapping population derived from two chickpea accessions having contrasting LRCs was genotyped by sequencing, and phenotyped to map four major quantitative trait loci (QTLs) contributing to 13-32% of the LRC trait variation. A single- nucleotide polymorphism tightly linked to the locus contributing to highest trait variation was located on the coding region of a gene (CaWIP2), orthologous to NO TRANSMITTING TRACT/WIP domain protein 2 (NTT/WIP2) gene of Arabidopsis thaliana. A polymorphic simple sequence repeat (SSR) in the CaWIP2 promoter showed differentiation between low versus high LRC parents and mapping individuals, suggesting its utility for marker-assisted selection. CaWIP2 promoter showed strong expression in chickpea apical root meristem and lateral root primordia. Expression of CaWIP2 under its native promoter in the Arabidopsis wip2wip4wip5 mutant rescued its rootless phenotype to produce more lateral roots than the wild-type plants, and led to formation of amyloplasts in the columella. CaWIP2 expression also induced the expression of genes that regulate lateral root emergence. Our study identified a gene-based marker for LRC which will be useful for developing drought-tolerant, high-yielding chickpea varieties.

Molecular mapping of two novel cold resistance genes in common wheat by 660K SNP array.

Low temperature and cold damage are natural factors that seriously reduce wheat yield. Thus, how to improve the cold resistance of wheat has been the focus of wheat breeders and geneticists. However, the genetic improvement for this trait has been slow, mainly because cold resistance is a complex quantitative trait and field phenotypic identification is relatively difficult. Therefore, the discovery, mapping, and cloning of the cold resistance genes of wheat provide a theoretical basis for the genetic improvement of wheat against cold resistance and facilitate the analysis of the molecular mechanisms of cold resistance in wheat. This study used the wheat line H261 and its EMS mutants LF2099 and XiNong 239 as materials. Cold trait segregation occurred in the F2 generation of mutants LF2099 and XiNong 239 at a 15:1 separation ratio. Genetic analysis showed that two dominant overlapping genes, temporarily named Wcr-3 and Wcr-4, control cold resistance in wheat. Furthermore, a combined BSA and SNP array established that Wcr-3 is between BU100519 (SSR marker) and AX-94843669 (SNP marker). The markers are 1.32 cM apart, corresponding to the 5.41 Mb physical interval on the Chinese Spring 2B chromosome with 67 functionally annotated genes. Wcr-4 is located between AX-94657955 (SNP marker) and LC-23 (SSR marker), which are 1.79 cM apart, corresponding to a 2.35 Mb physical interval on the Chinese Spring 2D chromosome, which contains 66 functionally annotated genes. Wcr-3 and Wcr-4 are two new cold resistance genes, laying the foundation for their fine mapping and cloning. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01425-w.

A simple sequence repeats marker of disease resistance in shrimp Litopenaeus vannamei and its application in selective breeding.

The polymorphism of the simple sequence repeat (SSR) in the 5' untranslated coding region (5'-UTR) of the antiviral gene IRF (LvIRF) has been shown to be implicated in the resistance to viral pathogens in shrimp Litopenaeus vannamei (L. vannamei). In this study, we explored the potential of this (CT)n-SSR marker in disease resistance breeding and the hereditary property of disease resistance traits in offspring. From 2018 to 2021, eight populations were generated through crossbreeding by selecting individuals according to microsatellite genotyping. Our results demonstrated that shrimp with the shorter (CT)n repeat exhibited higher resistance to white spot syndrome virus (WSSV) or Decapod iridescent virus 1 (DIV1); meanwhile, these resistance traits could be inherited in offspring. Interestingly, we observed that the longer (CT)n repeats were associated with bacterial resistance traits. Accordingly, shrimp with longer (CT)n repeats exhibited higher tolerance to Vibrio parahaemolyticus infection. Taken together, these results indicate that the single (CT)n-SSR marker could be used to selective breeding for both resistance to virus and bacteria in shrimps.

Genetic Diversity and Population Structure of Juniperus seravschanica Kom. Collected in Central Asia.

Juniperus seravschanica Kom. is a species that grows widely in the mountain ranges from Central Asia to Oman. It is an important tree for the formation of shrub-forest massifs in mountainous areas and for draining and fixing soils from middle to high altitudes. A comprehensive study of the species' genetic diversity and population structure is a basic approach to understanding the current status of J. seravschanica resources for the development of future conservation strategies. Samples from 15 populations of J. seravschanica were collected from the mountain ranges of Uzbekistan, Kyrgyzstan, and Kazakhstan. The genetic diversity and population structure of 15 Central Asian populations of J. seravschanica were assessed using 11 polymorphic simple sequence repeat (SSR) markers. Genetic diversity parameters, including the number of alleles (na), the effective number of alleles (ne), Shannon's information index (I), the percentage of polymorphic loci (PPL), Nei's genetic diversity index (Nei), principal coordinate analysis (PCoA), etc., were evaluated. The analysis of 15 J. seravschanica populations based on 11 polymorphic SSRs detected 35 alleles. The average PIC value was 0.432, and the highest value (0.662) was found in the JT_40 marker. Nei's genetic diversity index for the J. seravschanica populations was 0.450, ranging from 0.407 (population 14) to 0.566 (population 4). The analysis of molecular variance (AMOVA) showed that 90.3% of total genetic variation is distributed within the population. Using the alleles of all the populations, the gene flow (Nm) was found to be 4.654. Population structure analysis revealed poor clustering in the studied populations and confirmed our AMOVA results. The output of this work can be efficiently used for the maintenance of the species across the Central Asian region.

Multiple-Genome-Based Simple Sequence Repeat Is an Efficient and Successful Method in Genotyping and Classifying Different Jujube Germplasm Resources.

Jujube (Ziziphus jujuba Mill.) is a commercially important tree native to China, known for its high nutritional value and widespread distribution, as well as its diverse germplasm resources. Being resilient to harsh climatic conditions, the cultivation of jujube could provide a solution to food insecurity and income for people of arid and semi-arid regions in and outside of China. The evaluation of germplasm resources and genetic diversity in jujube necessitates the use of Simple Sequence Repeat (SSR) markers. SSR markers are highly polymorphic and can be used to evaluate the genetic diversity within and between cultivars of Chinese jujube, and are important for conservation biology, breeding programs, and the discovery of important traits for Chinese jujube improvement in China and abroad. However, traditional methods of SSR development are time-consuming and inadequate to meet the growing research demands. To address this issue, we developed a novel approach called Multiple-Genome-Based SSR identification (MGB-SSR), which utilizes the genomes of three jujube cultivars to rapidly screen for polymorphic SSRs in the jujube genome. Through the screening process, we identified 12 pairs of SSR primers, which were then used to successfully classify 249 jujube genotypes. Based on the genotyping results, a digital ID card was established, enabling the complete identification of all 249 jujube plants. The MGB-SSR approach proved efficient in rapidly detecting polymorphic SSRs within the jujube genome. Notably, this study represents the first successful differentiation of jujube germplasm resources using 12 SSR markers, with 4 markers successfully identifying triploid jujube genotypes. These findings offer valuable information for the classification of Chinese jujube germplasm, thereby providing significant assistance to jujube researchers and breeders in identifying unknown jujube germplasm.

Marker-Assisted Selection in Breeding for Fruit Trait Improvement: A Review.

Breeding fruit species is time-consuming and expensive. With few exceptions, trees are likely the worst species to work with in terms of genetics and breeding. Most are characterized by large trees, long juvenile periods, and intensive agricultural practice, and environmental variability plays an important role in the heritability evaluations of every single important trait. Although vegetative propagation allows for the production of a significant number of clonal replicates for the evaluation of environmental effects and genotype × environment interactions, the spaces required for plant cultivation and the intensity of work necessary for phenotypic surveys slow down the work of researchers. Fruit breeders are very often interested in fruit traits: size, weight, sugar and acid content, ripening time, fruit storability, and post-harvest practices, among other traits relevant to each individual species. The translation of trait loci and whole-genome sequences into diagnostic genetic markers that are effective and affordable for use by breeders, who must choose genetically superior parents and subsequently choose genetically superior individuals among their progeny, is one of the most difficult tasks still facing tree fruit geneticists. The availability of updated sequencing techniques and powerful software tools offered the opportunity to mine tens of fruit genomes to find out sequence variants potentially useful as molecular markers. This review is devoted to analysing what has been the role of molecular markers in assisting breeders in selection processes, with an emphasis on the fruit traits of the most important fruit crops for which examples of trustworthy molecular markers have been developed, such as the MDo.chr9.4 marker for red skin colour in apples, the CCD4-based marker CPRFC1, and LG3_13.146 marker for flesh colour in peaches, papayas, and cherries, respectively.

Pattern and variation in simple sequence repeat (SSR) at different genomic regions and its implications to maize evolution and breeding.

BACKGROUND: Repetitive DNA sequences accounts for over 80% of maize genome. Although simple sequence repeats (SSRs) account for only 0.03% of the genome, they have been widely used in maize genetic research and breeding as highly informative codominant DNA markers. The genome-wide distribution and polymorphism of SSRs are not well studied due to the lack of high-quality genome DNA sequence data. RESULTS: In this study, using data from high-quality de novo-sequenced maize genomes of five representative maize inbred lines, we revealed that SSRs were more densely present in telomeric region than centromeric region, and were more abundant in genic sequences than intergenic sequences. On genic sequences, tri- and hexanucleotide motifs were more abundant in CDS sequence and some mono- and dinucleotide motifs were more abundant in UTR sequences. Median length and chromosomal density of SSRs were both narrowly range-bound, with median length of 14-18 bp and genome-wide average density of 3355.77 bp/Mbp. LTR-RTs of < 0.4 Mya had higher SSR density (4498-4992 bp/Mbp). The genome-specific and motif-specific SSR polymorphism were studied. Their potential breeding applications were discussed. CONCLUSIONS: We found that the median length of SSR sequences of different SSR motifs was nearly constant. SSR density in genic regions was much higher than intergenic regions. In addition, SSR density at LTR-RTs of different evolutionary ages varied in a narrow range. The SSRs and their LTR-RT carriers evolved at an equal rate. All these observations indicated that SSR length and density were under control of yet unknown evolutionary forces. The chromosome region-specific and motif-specific SSR polymorphisms we observed supported the notion that SSR polymorphism was invaluable genome resource for developing highly informative genome and gene markers in maize genetic research and molecular breeding.

Development of genomic resources for Wenchengia alternifolia (Lamiaceae) based on genome skimming data.

Wenchengia alternifolia (Lamiaceae), the sole species of the genus Wenchengia is extremely rare and is currently listed as Critically Endangered (CR) on the IUCN Red List. The species had long been considered endemic to Hainan Island, China and was once believed to be extinct until a small remnant population was rediscovered at the type locality in 2010. Four more populations were later found on Hainan and in Vietnam. In order to develop genomic resources for further studies on population genetics and conservation biology of this rare species, we identified infraspecific molecular markers in the present study, using genome skimming data of five individuals collected from two populations on Hainan Island and three populations in Vietnam respectively. The length of plastome of the five individuals varied from 152,961 bp to 150,204 bp, and exhibited a typical angiosperm quadripartite structure. Six plastid hotspot regions with the Pi > 0.01 (trnH-psbA, psbA-trnK, rpl22, ndhE, ndhG-ndhI and rps15-ycf1), 1621 polymorphic gSSRs, as well as 1657 candidate SNPs in 237 variant nuclear genes were identified, thereby providing important information for further genetic studies.

Genetic Analysis of Mutagenesis That Induces the Photoperiod Insensitivity of Wild Cotton Gossypium hirsutum Subsp. purpurascens.

Cotton genus Gossypium L., especially its wild species, is rich in genetic diversity. However, this valuable genetic resource is barely used in cotton breeding programs. In part, due to photoperiod sensitivities, the genetic diversity of Gossypium remains largely untapped. Herein, we present a genetic analysis of morphological, cytological, and genomic changes from radiation-mediated mutagenesis that induced plant photoperiod insensitivity in the wild cotton of Gossypium hirsutum. Several morphological and agronomical traits were found to be highly inheritable using the progeny between the wild-type G. hirsutum subsp. purpurascens (El-Salvador) and its mutant line (Kupaysin). An analysis of pollen mother cells (PMCs) revealed quadrivalents that had an open ring shape and an adjoining type of divergence of chromosomes from translocation complexes. Using 336 SSR markers and 157 F2 progenies that were grown with parental genotypes and F1 hybrids in long day and short night conditions, five quantitative trait loci (QTLs) associated with cotton flowering were located on chromosomes At-05, At-11, and Dt-07. Nineteen candidate genes related to the flowering traits were suggested through molecular and in silico analysis. The DNA markers associated with the candidate genes, upon future functional analysis, would provide useful tools in marker-assisted selection (MAS) in cotton breeding programs for early flowering and maturity.

Classification of Southeast Asian mints (Mentha spp.) based on simple sequence repeat markers.

Mentha is a complex genus encompassing many species as a consequence of their interspecific hybridization and polyploidy. Southeast Asian mints have been poorly distinguished though they are widely used for culinary and medical purposes. In this study, we have analyzed Southeast Asian mints and known varieties as well as a related Lamiaceae species (Nepeta sp.) using simple sequence repeat (SSR) markers and leaf morphology. Two types of mints were clearly distinguished based on their venation pattern and leaf shape index. We developed 12 SSR markers that allowed good amplification in the Mentha and another Lamiaceae species. In the SSR-based phylogram, the Mentha lines could be delimited into groups I-VI. The Southeast Asian mints divided into groups I and II, and the phylogram separated most of the available species, with groups I and II containing the known species M. × cordifolia and M. arvensis, respectively. The separation of the two groups was supported by a population structure analysis. The SSR markers developed in this study enabled the simultaneous classification of mints and will help improve our understanding of the genetic composition of known mint varieties and as yet unclassified Southeast Asian mints.

Structured Populations of Critically Endangered Yellow Water Lily (Nuphar shimadai Hayata, Nymphaeaceae).

Yellow water lily (Nuphar shimadai Hayata) is a critically endangered species in Taiwan. Here, we examined genetic structures of four extant populations, WP, GPa, GPb and GPn, using 39 simple sequence repeat (SSR) markers. Positive genetic correlation was observed within 50 m, beyond which no correlation was detected due to isolation by distance according to Mantel correlogram. This suggests a significant genetic structuring of the species. Besides, multilocus genotype (MLG) analysis revealed that GPa was a panmictic population and the species' putative center of origin. Genetic exchange was observed between GPa and GPb populations, which likely resulted from their geographic proximity. Nevertheless, there was a strong asymmetric migration detected from GPa to WP, but a recent genetic barrier prevented dispersal further northward (WP). Geneland estimated the best number of clusters as K = 2, where WP distinctly separated from the rest of the populations. In STRUCTURE output of K = 3, a third cluster was abundant only in WP. We suggest to consider GPn and WP as separate conservation units, being far from GPa. There is indeed a need to investigate these populations; as predicted, Ne = 1.6 to 3.0 is considered low and that may put the species at risk of extinction.

High genetic diversity and low population differentiation of a medical plant Ficus hirta Vahl., uncovered by microsatellite loci: implications for conservation and breeding.

BACKGROUND: Wuzhimaotao (Radix Fici Hirtae) originates from the dry root of Ficus hirta (Moraceae), which is widely known as a medical and edible plant distributed in South China. As the increasing demand for Wuzhimaotao, the wild F. hirta has been extremely reduced during the past years. It is urgent to protect and rationally develop the wild resources of F. hirta for its sustainable utilization. However, a lack of genetic background of F. hirta makes it difficult to plan conservation and breeding strategies for this medical plant. In the present study, a total of 414 accessions of F. hirta from 7 provinces in southern China were evaluated for the population genetics using 9 polymorphic SSR markers. RESULTS: A mean of 17.1 alleles per locus was observed. The expected heterozygosity (He) varied from 0.142 to 0.861 (mean = 0.706) in nine SSR loci. High genetic diversity (He = 0.706, ranged from 0.613 to 0.755) and low genetic differentiation among populations (G'ST = 0.147) were revealed at population level. In addition, analysis of molecular variance (AMOVA) indicated that the principal molecular variance existed within populations (96.2%) was significantly higher than that among populations (3.8%). Meanwhile, the three kinds of clustering methods analysis (STRUCTURE, PCoA and UPGMA) suggested that the sampled populations were clustered into two main genetic groups (K = 2). Mantel test showed a significant correlation between geographic and genetic distance among populations (R2 = 0.281, P < 0.001). Pollen flow, seed flow and/or geographical barriers might be the main factors that formed the current genetic patterns of F. hirta populations. CONCLUSIONS: This is a comprehensive study of genetic diversity and population structure of F. hirta in southern China. We revealed the high genetic diversity and low population differentiation in this medicinal plant and clarified the causes of its current genetic patterns. Our study will provide novel insights into the exploitation and conservation strategies for F. hirta.

Association analysis of germination level cold stress tolerance and candidate gene identification in Upland cotton (Gossypium hirsutum L.).

Cotton originated from ancestors in the Gossypium genus that grew in semi-desert habitats. As a result, it is adversely affected by low temperatures especially during germination and the first weeks of growth. Despite this, there are relatively few molecular studies on cold stress in cotton. This limitation may present a future breeding handicap, as recent years have witnessed increased low temperature damage to cotton production. Cold tolerance is a sustainable approach to obtain good production in case of extreme cold. In the present study, 110 Upland cotton (Gossypium hirsutum) genotypes were evaluated for cold tolerance at the germination stage. We identified vigorous genotypes with cold-related parameters that outperformed the panel's average performance ( x ¯ = 76.9% CG, 83.9% CSI, 167.5 CWVI). Molecular genetic diversity analysis with 101 simple sequence repeat (SSR) markers yielding 416 loci was used to select tolerant genotypes that could be important materials for breeding this trait. A total of 16 marker-cold tolerance trait associations (p < 0.005) were identified with 10 of them having major effects (PVE > 10%). Based on the positions of these markers, candidate genes for cold tolerance in the G. hirsutum genome were identified. Three of these markers (BNL0569, CIR081 and CIR202) are important candidates for use in marker-assisted breeding for cold tolerance because they mapped to genes previously associated with cold tolerance in other plant species such as Arabidopsis thaliana, rice and tomato. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-022-01184-6.

Genetic Diversity Study on Geographical Populations of the Multipurpose Species Elsholtzia stauntonii Using Transferable Microsatellite Markers.

Elsholtzia stauntonii Benth. (Lamiaceae) is an economically important ornamental, medicinal and aromatic plant species. To meet the increasing market demand for E. stauntonii, it is necessary to assess genetic diversity within the species to accelerate the process of genetic improvement. Analysis of the transferability of simple sequence repeat (SSR) markers from related species or genera is a fast and economical method to evaluate diversity, and can ensure the availability of molecular markers in crops with limited genomic resources. In this study, the cross-genera transferability of 497 SSR markers selected from other members of the Lamiaceae (Salvia L., Perilla L., Mentha L., Hyptis Jacq., Leonurus L., Pogostemon Desf., Rosmarinus L., and Scutella L.) to E. stauntonii was 9.05% (45 primers). Among the 45 transferable markers, 10 markers revealed relatively high polymorphism in E. stauntonii. The genetic variation among 825 individuals from 18 natural populations of E. stauntonii in Hebei Province of China was analyzed using the 10 polymorphic SSR markers. On the basis of the SSR data, the average number of alleles (N A), expected heterozygosity (H E), and Shannon's information index (I) of the 10 primers pairs were 7.000, 0.478, and 0.688, respectively. Lower gene flow (N m = 1.252) and high genetic differentiation (F st = 0.181) were detected in the populations. Analysis of molecular variance (AMOVA) revealed that most of the variation (81.47%) was within the populations. Integrating the results of STRUCTURE, UPGMA (Unweighted Pair Group Method with Arithmetic Mean) clustering, and principal coordinate analysis, the 825 samples were grouped into two clusters associated with geographical provenance (southwestern and northeastern regions), which was consistent with the results of a Mantel test (r = 0.56, p < 0.001). Overall, SSR markers developed in related genera were effective to study the genetic structure and genetic diversity in geographical populations of E. stauntonii. The results provide a theoretical basis for conservation of genetic resources, genetic improvement, and construction of a core collection for E. stauntonii.

Analysis of Genetic Diversity of Banana Weevils (Cosmopolites sordidus) (Coleoptera: Curculionidae) Using Transcriptome-Derived Simple Sequence Repeat Markers.

The banana weevil, Cosmopolites sordidus (Germar) (Coleoptera: Curculionidae) is an economically important insect pest of bananas. It causes up to 100% yield losses and substantial lifespan reduction in bananas. Advances in genomics, proteomics, and sequencing technologies have provided powerful pathways to genotyping disastrous pests such as C. sordidus. However, such technologies are often not available to the majority of rural subtropical African banana growers and pest control managers. This study was therefore motivated by the need to create cheap and easily accessible C. sordidus genotyping methods that could be deployed by banana pest control managers to the benefit of C. sordidus control programs in the tropics where such advanced technologies are not readily accessible. We used an in-house C. sordidus transcriptome from the an-ongoing study from which we mined an array of simple sequence repeat (SSR) markers. Of these, six highly polymorphic transcriptome-derived SSR markers were used to successfully genotype within and among banana weevil population genetic diversity of 12 C. sordidus populations collected from four banana-growing agro-ecological zones (AEZs) in Uganda. The developed transcriptome-derived SSR markers can be used by researchers in population genetics for characterization of the C. sordidus and identification of new genes that are linked to traits of particular interest. The significant genetic diversity revealed in C. sordidus provides pertinent information for integrated pest management strategies.

Full-Length SMRT Transcriptome Sequencing and SSR Analysis of Bactrocera dorsalis (Hendel).

Bactrocera dorsalis (Hendel), as one of the most notorious and destructive invasive agricultural pests in the world, causes damage to over 250 different types of fruits and vegetables throughout tropical and subtropical areas. PacBio single-molecule real-time (SMRT) sequencing was used to generate the full-length transcriptome data of B. dorsalis. A total of 40,319,890 subreads (76.6 Gb, clean reads) were generated, including 535,241 circular consensus sequences (CCSs) and 386,916 full-length non-concatemer reads (FLNCs). Transcript cluster analysis of the FLNC reads revealed 22,780 high-quality reads (HQs). In total, 12,274 transcripts were functionally annotated based on four different databases. A total of 1978 SSR loci were distributed throughout 1714 HQ transcripts, of which 1926 were complete SSRs and 52 were complex SSRs. Among the total SSR loci, 2-3 nucleotide repeats were dominant, occupying 83.62%, of which di- and tri- nucleotide repeats were 39.38% and 44.24%, respectively. We detected 105 repeat motifs, of which AT/AT (50.19%), AC/GT (39.15%), CAA/TTG (32.46%), and ACA/TGT (10.86%) were the most common in di- and tri-nucleotide repeats. The repeat SSR motifs were 12-190 bp in length, and 1638 (88.02%) were shorter than 20 bp. According to the randomly selected microsatellite sequence, 80 pairs of primers were designed, and 174 individuals were randomly amplified by PCR using primers. The number of primers that had amplification products with clear bands and showed good polymorphism came to 41, indicating that this was a feasible way to explore SSR markers from the transcriptomic data of B. dorsalis. These results lay a foundation for developing highly polymorphic microsatellites for researching the functional genomics, population genetic structure, and genetic diversity of B. dorsalis.

Haplotype and diversity analysis of indigenous rice for salinity tolerance in early-stage seedling using simple sequence repeat markers.

We evaluated the seedling-stage salt tolerance of a total of 50 indigenous rice genotypes from coastal Tamil Nadu. Using a hydroponic system, we studied the different agronomic characters 14 days after exposure to six different concentrations of saline solution. Shoot and root length as well as plant biomass at seedling stage decreased with increasing salinity. Genotypes showing significant interaction and differential response towards salinity were assessed at the molecular level using 20 simple sequence repeat (SSR) markers linked with salt-tolerance QTL. These genotypes were grouped into eleven clusters based on molecular diversity analysis and eight clusters based on D2 statistical analysis. We found wide genetic distance among the genotypes studied. Simple correlation analysis revealed highly significant associations among the traits studied. The combination of morphological findings and molecular assessment revealed better salt-tolerance in a few genotypes viz. Kuzhi adichan, Poonkar, Kallundai, and Sornamugi.