High Polymorphism Information Content Research Articles

Genetic diversity of eucalypts for germplasm conservation in Forest Area with the Special Purpose of Mount Bromo, Karanganyar, Indonesia

Abstract. Rahayu, Fatimah, Wiwoho J, Firdaus SU, Pujiyono, Marimin, Arianto DP, Pramono A. 2021. Genetic diversity of eucalypts for germplasm conservation in Forest Area with the Special Purpose of Mount Bromo, Karanganyar, Indonesia. Biodiversitas 22: 4223-4235. As a repository of a gene pool, eucalypts germplasm enriches biodiversity, maintains ecosystem sustainability, and aids in conservation. Therefore, this study aims to analyze the genetic diversity of eucalypts (Corymbia and Eucalyptus) for the development of germplasm conservation in Forest Area with the Special Purpose (KHDTK) Bromo Forest, Karanganyar, Indonesia. In this study, 14 simple sequence repeat (SSR) markers were used to assess the genetic diversity among 20 accessions (Corymbia and 5 Eucalyptus species) from Central and West Java. Subsequently, the genetic parameters were measured and a phylogenetic tree was constructed. The result showed that the SSR markers have high variability, although they belong to different genera. Furthermore, the genetic diversity showed 49 alleles with an average of 3 alleles per locus, while the polymorphism information content (PIC) values were 0.55. There were 4 SSR markers (EMBRA13, EMBRA8, EMCRC11, and EMBRA2) with high PIC value, while the gene diversity (He) of Corymbia and 5 Eucalyptus showed a low level of genetic diversity. The genetic relationship and population structure were divided into genera Corymbia and Eucalyptus. For further application, the eucalypt cultivated in the KHDTK Bromo Forest can contribute as a reference set and 14 SSR markers as a potential marker in combination with morphological characterization to generate a database for germplasm management and conservation.

Assessment of Genetic Variability among Onion (Allium cepa L.) Cultivars using RAPD and SRAP Molecular Markers

Onion (Allium cepa L.) is one of the most important vegetable crops worldwide, however, limited studies are available on genetic variability of onion resources. The aim of this study was to assess the genetic diversity of twelve onion cultivars which are widely grown in different regions of Sudan, using RAPD and SRAP markers. A total of 32 RAPD and 21 SRAPprimers were screened. To assess variability, the data generated were used to group the cultivars using Jaccard’s coefficient of similarity. Only 20, out of tested RAPD primers were producible and showed 326 different-sized DNAfragments with one or more of the tested cultivars giving an average of 27.17 alleles per cultivar. Hundred percent polymorphism was recorded for each of the primers tested with high polymorphism information content (PIC) rangingfrom 0.89 to 1.00. In SRAP analysis, 8 out of 21 primer pairs tested, produced bands of different sizes. A total of 66 DNA fragments were detected for the 12 cultivars with an average of 5.5 alleles per cultivar. The number of different fragments generated by each primer ranged from one to eight and 100 percent polymorphism was recorded for each of the primerstested with high PIC of approximately 1.00. Similarity matrices constructed using RAPD, SRAP or the combined data suggested a relatively high level of genetic diversity among the 12 onion cultivars, the most diverse cultivars were Shendi yellow and Baftaim red hybrid with 14 % similarity. The study revealed that although both molecular markers used were efficient inelucidating genetic diversity among the tested onion cultivars, SRAP markers were the most potential and could be used as an integrated approach in onion breeding programmes.

Association analysis, genetic diversity and population structure of barley (Hordeum vulgare L.) under heat stress conditions using SSR and ISSR markers linked to primary and secondary metabolites.

Barley is one of the major cereal crops, which can provide a significant source of genes for stress tolerance due to its high diversity and adaptability. Metabolite traits are considered to be significant for adaptation of barley to heat stress. In the present study, genetic relationships between 120 barley genotypes were determined with 50 simple sequence repeat (SSR) and 26 inter simple sequence repeat (ISSR) markers under heat stress and non-stress conditions. Moreover, genetic diversity of barley accessions was investigated using the studied markers covering 7 chromosomes of barley. In general, 153 and 85 polymorphic alleles were detected for SSR and ISSR and number of the observed polymorphic allele varied between 2-9 and 2-6, with an average of 3.26 and 3.26 alleles per locus, respectively. Markers of Bmag0223, GBMS180/180, HVM7, ISSR22, ISSR25, and ISSR48 were the most informative due to their high polymorphism information content value demonstrating that putative techniques utilized in this research can be powerful and valuable tools in breeding program of barley. Association analysis was performed between 9 important traits and SSR and ISSR markers using four statistical models. The results revealed that the model containing both population structure (Q) and general similarity in genetic background arising from shared kinship (K) factors reduced false positive associations between markers and phenotypes. According to the results, some of markers related to more than one trait under normal conditions (ISSR31-2, HVM62, and GBMS180/180) and heat stress conditions (ISSR20-5, EBmac635, HVM14, and ISSR37-3) were determined, which can be considered to be the most interesting candidates for further studies and simultaneously will provide a useful target for the future breeding programs, such as marker-assisted selection (MAS).

Trends in the molecular epidemiology and population genetics of emerging Sporothrix species.

Sporothrix (Ophiostomatales) comprises species that are pathogenic to humans and other mammals as well as environmental fungi. Developments in molecular phylogeny have changed our perceptions about the epidemiology, host-association, and virulence of Sporothrix. The classical agent of sporotrichosis, Sporothrix schenckii, now comprises several species nested in a clinical clade with S. brasiliensis, S. globosa, and S. luriei. To gain a more precise view of outbreaks dynamics, structure, and origin of genetic variation within and among populations of Sporothrix, we applied three sets of discriminatory AFLP markers (#3 EcoRI-GA/MseI-TT, #5 EcoRI-GA/MseI-AG, and #6 EcoRI-TA/MseI-AA) and mating-type analysis to a large collection of human, animal and environmental isolates spanning the major endemic areas. A total of 451 polymorphic loci were amplified in vitro from 188 samples, and revealed high polymorphism information content (PIC = 0.1765–0.2253), marker index (MI = 0.0001–0.0002), effective multiplex ratio (E = 15.1720–23.5591), resolving power (Rp = 26.1075–40.2795), discriminating power (D = 0.9766–0.9879), expected heterozygosity (H = 0.1957–0.2588), and mean heterozygosity (Havp = 0.000007–0.000009), demonstrating the effectiveness of AFLP markers to speciate Sporothrix. Analysis using the program structure indicated three genetic clusters matching S. brasiliensis (population 1), S. schenckii (population 2), and S. globosa (population 3), with the presence of patterns of admixture amongst all populations. AMOVA revealed highly structured clusters (PhiPT = 0.458–0.484, P < 0.0001), with roughly equivalent genetic variability within (46–48 %) and between (52–54 %) populations. Heterothallism was the exclusive mating strategy, and the distributions of MAT1-1 or MAT1-2 idiomorphs were not significantly skewed (1:1 ratio) for S. schenckii (χ2 = 2.522; P = 0.1122), supporting random mating. In contrast, skewed distributions were found for S. globosa (χ2 = 9.529; P = 0.0020) with a predominance of MAT1-1 isolates, and regional differences were highlighted for S. brasiliensis with the overwhelming occurrence of MAT1-2 in Rio de Janeiro (χ2 = 14.222; P = 0.0002) and Pernambuco (χ2 = 7.364; P = 0.0067), in comparison to a higher prevalence of MAT1-1 in the Rio Grande do Sul (χ2 = 7.364; P = 0.0067). Epidemiological trends reveal the geographic expansion of cat-transmitted sporotrichosis due to S. brasiliensis via founder effect. These data support Rio de Janeiro as the centre of origin that has led to the spread of this disease to other regions in Brazil. Our ability to reconstruct the source, spread, and evolution of the ongoing outbreaks from molecular data provides high-quality information for decision-making aimed at mitigating the progression of the disease. Other uses include surveillance, rapid diagnosis, case connectivity, and guiding access to appropriate antifungal treatment.

Assessment of the Genetic Diversity among Wild Colocynth (Citrullus colocynthis (L.)Schrad.) Ecotypes Collected from Different Regions of Sudan

Better utilization and conservation of the medicinal and aromatic plants (MAP), most of which are collected from the wild, necessitate their screening to identify potential genotypes for both production and utilization in interbreeding to develop superior genotypes.The objective of this research, therefore,wasthe assessment of the genetic relationship among wild colocynth Citrullus colocynthis (L.)Schrad. ecotypes. Seeds of twelve colocynth plants (SF, K10, K101, K114, ABD, SH, G, GD, GD1, ND, SD and NK) were collected from wild plants growing in twelve diverse ecologies of Sudan’s States. Commercial watermelon (Wm) seeds were obtained from the seed store and tested for comparison. Randomly Amplified Polymorphic DNA (RAPD) technology was used to explore the genetic relatedness of the ecotypes. Ten RAPD primers were used to amplify CTAB-extracted DNA from leaves of each ecotype. Data were used to create similarity matrices using the PAST 3.01 software package and a dendrogram was constructed based on Jaccard’s similarity coefficients. A total of 321 DNA fragments were detected by RAPD markers for the 13 tested plants giving an average of 24.69 alleles per ecotype. The number of fragments detected for each ecotype ranged from 11 (GD) to 64 detected for K10; ten fragments were detected for watermelon. Hundred percent polymorphism was recorded for the tested primers with high Polymorphism Information Content (PIC) values in the range of 0.97-1.00. Similarity matrix constructed from data generated indicated that the closest ecotypes, with 67% similarity, were ABD and K114, while the most distant ones (no similarity) were SD and GD as well as GD and Wm. The constructed dendrogram divided the 12 colocynth ecotypes and the watermelon cultivar into three major clusters; the first includes watermelon cultivar as an out-group, the second includes all ecotypes except K10 which is grouped in a separate cluster. These results conclude that the ecological zone does not affect the genetic relatedness of the tested ecotypes despite their high genetic polymorphism. The variability between the ecotypes points out that Sudan’s colocynth germplasm possesses high potentiality for future improvement through crossing and selection.

Development and Characterization of Novel Polymorphic Microsatellite Markers for Tapinoma indicum (Hymenoptera: Formicidae).

Tapinoma indicum (Forel) (Hymenoptera: Formicidae) is a nuisance pest in Asia countries. However, studies on T. indicum are limited, especially in the field of molecular biology, to investigate the species characteristic at the molecular level. This paper aims to provide valuable genetic markers as tools with which to study the T. indicum population. In this study, a total of 143,998 microsatellite markers were developed based on the 2.61 × 106 microsatellites isolated from T. indicum genomic DNA sequences. Fifty selected microsatellite markers were amplified with varying numbers of alleles ranging from 0 to 19. Seven out of fifty microsatellite markers were characterized for polymorphism with the Hardy–Weinberg equilibrium (HWE) and linkage disequilibrium (LD) analysis. All seven microsatellite markers demonstrated a high polymorphic information content (PIC) value ranging from 0.87 to 0.93, with a mean value of 0.90. There is no evidence of scoring errors caused by stutter peaks, no large allele dropout, and no linkage disequilibrium among the seven loci; although loci Ti-Tr04, Ti-Tr09, Ti-Te04, Ti-Te13, and Ti-Pe5 showed signs of null alleles and deviation from the HWE due to excessive homozygosity. In conclusion, a significant amount of microsatellite markers was developed from the data set of next-generation sequencing, and seven of microsatellite markers were validated as informative genetic markers that can be utilized to study the T. indicum population.

Genetic diversity of local rice varieties (Oryza sativa L.) in Vietnam’s Mekong Delta based on SSR markers and morphological characteristics

Based on target traits, use of the genetic diversity of rice is beneficial for crop improvement. In this study, 41 rice varieties local to Vietnam’s Mekong Delta were evaluated on the basis of 11 quantitative morphological traits, along with the assessment of genetic diversity according to 50 SSR markers. The actual yield had a significance level of 0.05, while plant height and panicles per square meter had a high significance level of 0.001. Cluster analysis based on 11 quantitative traits also revealed that two were the optimal number of clusters used in this study. The highest polymorphic information content (PIC) value obtained was for RM286 (0.49), with a range of 0.00 to 0.49 and an average PIC of 0.14. Both structure and phylogenetic tree analyses as inferred from 50 SSR markers by the unweighted pair‐group method with arithmetic mean (UPGMA) also indicated that the 41 local rice varieties could be divided into two major groups. This study provides a useful information for Mot bui do cao CM, and Mot bui five varieties for improvements in the yield and intermediate amylose content of local rice‐breeding programs in future, especially for the Mekong Delta region.

Genetic Characterization of Brassica oleracea L. var. acephala DC cultivars from Bosnia and Herzegovina

&#x0D; &#x0D; Fifteen Brassica oleracea L. var. acephala DC accessions were collected and evaluated for their genetic diversity and molecular characteristics using 10 previously developed and used codominant SSR markers. The SSR loci used revealed a total of 51 alleles. The domestic kale varieties were collected locally and the hybrids kales were found in markets. The averages of total heterozygosity (H) and polymorphic information content (PIC) was 0.6774 and 0.6201, respectively. The average number of alleles was 4.2 per locus, where 8 of 10 markers showed high polymorphic information content (PIC), being more than 0.5. The genetic relationships among the populations revealed by Unweighted Pair Group Method with Arithmetic mean (UPGMA), showed a clear clustering into three distinct groups. As expected, all local varieties were separated within two separated clusters, whereas only two domestic varieties shared one cluster with the group of hybrids, indicating similar genetic background shared by domestic and hybrid varieties. These results indicate and confirm the regional and physiological differences between the kale varieties and confirm the SSR marker efficiency for genetic characterization assays&#x0D; &#x0D; &#x0D; &#x0D; &#x0D;

Assessment of genetic diversity using microsatellite markers to compare donkeys (Equus asinus) with horses (Equus caballus).

ObjectiveThe study aimed to evaluate the diversity of donkey populations by comparing with the diversity of Thoroughbred and Jeju Halla horses; identified breeding backgrounds can contribute to management and conservation of donkeys in South Korea.MethodsA total of 100 horse (50 Thoroughbreds and 50 Jeju Halla horses) and 79 donkeys samples were genotyped with 15 microsatellite markers (AHT4, AHT5, ASB2, ASB17, ASB23, CA425, HMS1, HMS2, HMS3, HMS6, HMS7, HTG4, HTG10, LEX3, and VHL20), to identify genetic diversity and relationships among horses and donkeys.ResultsThe observed number of alleles per locus ranged from 1 (ASB17, HMS1) to 14 (AHT5), with a mean value of 4.87, 8.00, and 5.87 in Thoroughbreds, Jeju Halla horses, and donkeys, respectively. Of the 15 markers, AHT4, AHT5, ASB23, CA425, HMS2, HMS3, HTG4, HTG10, and LEX3 loci had relatively high polymorphism information content (PIC) values (PIC>0.5) in these three populations. Mean levels of genetic variation were HE = 0.6721 and HO = 0.6600 in Thoroughbreds, HE = 0.7898 and HO = 0.7100 in Jeju Halla horses, and HE = 0.5635 and HO = 0.4861 in donkeys. Of the 15 loci in donkeys, three loci had negative inbreeding coefficients (FIS), with a moderate mean FIS (0.138). The FIS estimate for the HTG4 marker was highest (0.531) and HMS6 marker was lowest (−0.001). The total probability of exclusion value of 15 microsatellite loci was 0.9996 in donkeys.ConclusionGenetic cluster analysis showed that the genetic relationship among 79 donkeys was generally consistent with pedigree records. Among the three breeds, donkeys and Thoroughbred horses formed clearly different groups, but the group of Jeju Halla horses overlapped with that of Thoroughbred horses, suggesting that the loci would be suitable for donkey parentage testing. Therefore, the results of this study are a valid tool for genetic study and conservation of donkeys.

Genetic Diversity and Population Assessment of Musa L. (Musaceae) Employing CDDP Markers

Sixty-six accessions of Musa genus with different genomic groups that consisted of wild relatives and cultivated lines were obtained from the International Transit Center, Belgium, for DNA extraction using Cetyl trimethylammonium bromide method, followed by amplification with Conserved DNA-derived Polymorphism (CDDP) markers for genetic diversity and population assessment. A total of 421 alleles with major allele frequency of 2.051 were detected from the reproducible markers. High genetic diversity (GD, 11.093) and polymorphic information content (0.918) were revealed. The number of polymorphic loci and percentage of polymorphic loci ranged from 59 to 66 and 89.34 to 100, respectively. Using the potential genetic indicators including effective number of alleles, Nei’s genetic diversity, and Shannon’s information index, the AS genomic group was identified to have the highest GD, while the AAA accessions had the lowest GD indices. The GD parameters identified in the accessions were ranked as AS > AAB > AAAA > AA > ABB > wild diploidy > BB > AB > AAA from high to low based on polymorphic loci of the markers. Total intraspecific GD, interspecific GD, and estimate gene flow identified were 0.433, 0.404, and 7.113, respectively. The coefficient of gene differentiation of 0.066 was obtained, indicating 6.57% among the population and 93.43% within the population. Dendrogram analysis produced nine major groups with subgroups at similarity index of 0.814. These CDDP functional gene-based markers were informative and very efficient in resolving GD, and population indices among the banana and plantain accessions of different genomes. The identified CDDP markers might serve as potential tools for selecting suitable training populations for breeding and conservation of Musa species.

Genome-wide identification of microsatellite and development of polymorphic SSR markers for spotted sea bass (Lateolabrax maculatus)

The spotted sea bass (Lateolabrax maculatus) is a valuable commercial mariculture fish in China and has huge potential for future genetic improvement. However, only limited number of SSR (simple sequence repeat) markers have been developed for L. maculatus. In this study, a total of 220,709 SSRs were identified from the L. maculatus genome. The total length, density and frequency of SSRs were 5,119,809 bp, 9898.28 bp/Mb and 425.06 loci/Mb, respectively. Dinucleotide repeats were predominant among SSRs (85.00 %) and AC was the most abundant SSR motifs. Both the number and length of SSRs in each chromosome were positively associated with the chromosome length. A total of 155,905 SSR markers were successfully designed (with an average of 6496 SSRs per chromosome). The frequency of SSR markers varied from 245.63 per Mb to 346.60 per Mb. 14 polymorphic SSR primers were used to evaluate the genetic diversity of L. maculatus wild population from Zhuhai. A total of 74 alleles were detected and most SSR markers showed high polymorphism information content (PIC). The cross-species transferability of 11 out of 67 tested loci were proven successful in largemouth bass Micropterus salmoides. The genome-wide SSRs and newly developed SSR markers made available in this study will facilitate future genetic breeding and genomic studies of spotted sea bass.

Genetic diversity of improved genotypes of Tectona grandis in the state of Mato Grosso, Brazil

Teak (Tectona grandis) is a tree species native to South-east Asia and widely cultivated in Latin America. In Mato Grosso state, Brazil, teak is the second-most planted forest species due to its high productivity resulting from high adaptability to the soil and climate conditions of the region. However, there is little information about the genetic diversity of improved genotypes cultivated in the state. Thus, the objective of this study was to quantify, through ISSR (inter-simple sequence repeat) molecular markers, the genetic diversity of improved teak genotypes used in different plantations in Mato Grosso state. For this, 31 ISSR primers were tested on 113 genotypes separated into nine populations. Six primers were selected to perform the analyses, resulting in 55 amplified loci. All selected primers showed a high polymorphic information content. Considering all genotypes in the Mato Grosso population, the percentage of polymorphic loci (P) was 96.4%, the genetic diversity according to Nei՚s genetic diversity index (H) was 1.96, and according to Shannon՚s information/diversity index (I), it was 0.53. Considering each population individually, P varied from 50.9% to 90.9%, H from 0.21 to 0.33, and I from 0.30 to 0.49. Most of the genetic diversity observed was within populations (86%). Four genetic groups were identified, with a high level of admixture among them. The existence of a considerable genetic diversity within populations indicates the possibility of exploring this variation in genetic improvement programmes. However, the introduction of new genetic materials is recommended to increase the genetic base of this species in Brazil.

Genetic characterization of the orphan crop tef [Eragrostis tef (Zucc.) Trotter] accessions using simple sequence repeat markers

Tef is the most important and well adapted cereal crop in Ethiopia and grows better than other cereals both in biotic and abiotic stress conditions. In this study 93 tef accessions representative of various agro-ecologies of Ethiopia along with four improved varieties, one local check and six wild relatives were studied. The materials were genotyped with 12 simple sequence repeat markers (SSRs) to investigate the extent and structure of the genetic diversity of tef. A total of 152 alleles were detected indicating high polymorphism in the studied accessions. The number of alleles per polymorphic loci ranged from seven for CNLT 538 to 29 for CNLT150 with an average of 12.67. High average polymorphic information content (0.82) and average expected heterozygosity (0.86) also signifies the suitability and efficiency of SSR markers to discriminate among tef genotypes. Cluster, principal coordinate and structure analyses grouped the 98 tef genotypes and six wild relatives into four distinct classes. The groupings showed no special pattern despite the different regions of origins for the accessions; however, the five wild relatives were grouped together. The analysis of molecular variance and fixation index showed low genetic differentiation and high gene flow between regions revealing that much of the variability was attributed to within populations variation. Generally, the present study showed that there is high genetic variability with unstructured genetic pattern among the studied tef genotypes which implies the vitality of conserved tef germplasms at Ethiopian Biodiversity Institute and the impact of the seed exchange system on tef breeding activities in Ethiopia.

Revealing genetic variation in mini core germplasm of urdbean (Vigna mungo (L.) Hepper)

A set of 47 urdbean genotypes including 12 promising varieties and 34 popular land races and a popular OUAT variety “Ujala” (standard check) were characterized for genetic variation based on molecular markers and morpho-economic traits. The molecular marker- based genotyping revealed a tremendous higher level of polymorphism (97.05%) with high average PIC (polymorphic information content) value (0.75). ISSR 1357 was considered highly informative that revealed the highest PIC (0.87) and marker index value (MI:5.25). An 840 bp allele (band) was characteristic to Kantapada local - A, Kendrapada local-D, and Nayagarh local - C. Such genotype- specific finger-printing may serve for reliable varietal characterization and elimination of duplicates. The test genotypes were grouped into six distinct clusters. TU 10-13, LBG 623, TAU 1, OBG 33, LBG 17 and PU 31 were highly divergent. PU 31 had inherent high yield potential (˃5.0q/ha) with known YMV resistance. The above divergent high yielding test genotypes may serve as candidate varieties for further genetic improvement using recombination breeding.

Confirmation of hybridity in DOGR hybrids of onion (Allium cepa L.) using SSR markers

Molecular identification and hybrid purity testing in nineonion hybrids along with their seven parental lines wasconducted at ICAR-Directorate of Onion and GarlicResearch, Rajgurunagar, Pune (Maharashtra). Four out often microsatellite (SSRs- Simple Sequence Repeats) markerswere found to be polymorphic. All four polymorphic markersrevealed polymorphism between the male and female parentsof one or more hybrids and was utilized for their hybriditytesting. ACM016, ACM033 and ACM078 have beenidentified as the three diagnostic markers for testing thegenetic purity of all the nine onion hybrids included in thisstudy viz.; DOGR Hy-1, DOGR Hy-2, DOGR Hy-3, DOGRHy-4, DOGR Hy-5, DOGR Hy-6, DOGR Hy-7, DOGR Hy-8and DOGR Hy-50. These 10 SSR loci amplified a total 28alleles in 9 hybrids and 7 parents with 2-4 alleles per loci.The average number of alleles per loci was found to be 2.8.The highest polymorphism information content (PIC value)was observed to be 0.7137 for the marker ACM093 and thethree potential markers ACM016, ACM033 and ACM078with PIC value 0.5874, 0.5949 and 0.6628, respectively werethe best markers that could be used to differentiate theparental lines and utilized for hybridity testing. This studyshowed that SSR are reliable and effective molecular markercapable of detecting genetic purity of onion hybrids.

Short Communication: MADS-box genes expression in seedlings of Elaeis guineensis Jacq. resistance to Ganoderma boninense

Abstract. Hayati R, Lisnawita, Munir E, Basyuni M. 2020. Short Communication: MADS-box genes expression in seedlings of Elaeis guineensis Jacq. resistance to Ganoderma boninense. Biodiversitas 21: 5805-5810. Advances in molecular technology were allowed the molecular studies of the flowering genes to expected to be used to predict oil palm (Elaeis guineensis) fruit production. This study aims to identify the genetic structure associated with flowering genes, belonging to the MADS-box gene family of E. guineensis to Ganoderma boninense susceptible and resistance types. Different length of expression genes was identified for each population displayed the single band pattern. On the basis of UVITEC-1D methods, all populations (RA, RS, SS, and SA) detected overall expression in primer EGGLO and EGDEF. Moreover, several populations (SS, SA, RA) were not expressed in primer EGSQUA, EGAGL2, and EGAG. Based on the GENEALEX results, 5 sets of MADS-box family markers showed Na values were 2.20 to 3.00, and He values were 0.40–0.67. High polymorphism information content (PIC &gt; 0.50) was found in Resistance Asymptomatic (RA) and Susceptible Asymptomatic (SA) populations (PIC = 0.82). The phylogenetic tree described clustering for populations based on genetic distances. The dendrogram was constructed with two cluster groups. The first cluster was described as resistance asymptomatic and symptomatic population. Furthermore, the second cluster consisted of two clustering susceptible asymptomatic and symptomatic populations, separately. Several oil palm genes resistant symptomatic and asymptomatic was found in one cluster, similarly in susceptible symptomatic and asymptomatic grouped more closely. This phenomenon is likely to be the sampling rate and the addictive nature of parent crossing derivatives.&#x0D;

SNP marker assay and candidate gene identification for sex expression via genotyping-by-sequencing-based genome-wide associations (GWAS) analyses in Oriental melon (Cucumis melo L.var.makuwa)

The Oriental melon is an important delicious fruit crop across East Asia. Single-nucleotide polymorphisms (SNPs) are considered to be a useful genotyping tool both to detect genetic diversity and to protect breeder rights. This study used genotyping-by-sequencing (GBS) to detect genome-wide SNPs by analyzing 48 commercial Oriental melon varieties to generate 5640 filtered SNPs. Based on the high polymorphism information content (PIC), genetic and physical distances, a subset of 192 SNPs was selected as putative SNPs for validation via the Fluidigm JunoTM system. Of these, 164 SNPs were successfully validated in 87 Oriental melon varieties with a validation ratio of 85.41 %. Furthermore, these 87 Oriental melon varieties were classified into ten distinct groups, based on 164 SNP markers. Therefore, these large set of SNP markers detected here has several application, such as genetic diversity studies, varietal identification and marker-assisted selection (MAS) in the Oriental melon. Genome-wide associations (GWAS) analyses detected significant 18 SNPs associated with various morphological traits, including two novel SNPs for sex expression on chromosomes 1 and 8 that were not colocalized with previous studies. The four potential candidate genes such as MELO3C015898, MELO3C015904, MELO3C024563, and MELO3C024565 were predicted within the GWAS-SNPs regions for sex expression in the Oriental melon. Hence, identification of these candidate genes has provided the foundation to study the molecular genetic mechanism of sex expression trait in the Oriental melon.

Genetic diversity in pearl millet restorers based on seed vigour traits and SSR markers

Thirty four pearl millet restorer lines were evaluated on the basis of nine seed vigour traits and molecular analysis was done with 55 SSRs to study the genetic divergence among them. Cluster analysis based on seed vigour parameters revealed the considerable amount of variability and all genotypes were divided into six clusters. Furthermore, a set of 39 SSRs, selected after initial screening of 55, amplified 226 alleles with a mean of 5.84 alleles per locus. The highest polymorphic information content (PIC) value obtained was for PSMP 2084 (0.88) with a range of 0 to 0.88 and average PIC of 0.558. Neighbor joining tree based on molecular data differentiated all the genotypes into two major groups. The grouping based on both approaches can help breeders to use the studied material in further breeding for improvement of germination potential and early seedling vigour of pearl millet.

The landscape of microsatellites in the enset (Ensete ventricosum) genome and web-based marker resource development

Ensete ventricosum (Musaceae, enset) is an Ethiopian food security crop. To realize the potential of enset for rural livelihoods, further knowledge of enset diversity, genetics and genomics is required to support breeding programs and conservation. This study was conducted to explore the enset genome to develop molecular markers, genomics resources, and characterize enset landraces while giving insight into the organization of the genome. We identified 233 microsatellites (simple sequence repeats, SSRs) per Mbp in the enset genome, representing 0.28% of the genome. Mono- and di-nucleotide repeats motifs were found in a higher proportion than other classes of SSR-motifs. In total, 154,586 non-redundant enset microsatellite markers (EMM) were identified and 40 selected for primer development. Marker validation by PCR and low-cost agarose gel electrophoresis revealed that 92.5% were polymorphic, showing a high PIC (Polymorphism Information Content; 0.87) and expected heterozygosity (He = 0.79–0.82). In silico analysis of genomes of closely related species showed 46.86% of the markers were transferable among enset species and 1.90% were transferable to Musa. The SSRs are robust (with basic PCR methods and agarose gel electrophoresis), informative, and applicable in measuring enset diversity, genotyping, selection and potentially breeding. Enset SSRs are available in a web-based database at https://enset-project.org/EnMom@base.html (or https://enset.aau.edu.et/index.html, downloadable from Figshare).

Development and application of perfect SSR markers in cotton

BackgroundThis study aimed to develop a set of perfect simple sequence repeat (SSR) markers with a single copy in the cotton genome, to construct a DNA fingerprint database suitable for authentication of cotton cultivars. We optimized the polymerase chain reaction (PCR) system for multi-platform compatibility and improving detection efficiency. Based on the reference genome of upland cotton and 10× resequencing data of 48 basic cotton germplasm lines, single-copy polymorphic SSR sites were identified and developed as diploidization SSR markers. The SSR markers were detected by denaturing polyacrylamide gel electrophoresis (PAGE) for initial screening, then fluorescence capillary electrophoresis for secondary screening. The final perfect SSR markers were evaluated and verified using 210 lines from different sources among Chinese cotton regional trials.ResultsUsing bioinformatics techniques, 1 246 SSR markers were designed from 26 626 single-copy SSR loci. Adopting a stepwise (primary and secondary) screening strategy, a set of 60 perfect SSR markers was selected with high amplification efficiency and stability, easy interpretation of peak type, multiple allelic variations, high polymorphism information content (PIC) value, uniform chromosome distribution, and single-copy characteristics. A multiplex PCR system was established with ten SSR markers using capillary electrophoresis detection.ConclusionsA set of perfect SSR markers of cotton was developed and a high-throughput SSR marker detection system was established. This study lays a foundation for large-scale and standardized construction of a cotton DNA fingerprint database for authentication of cotton varieties.