Morphological and molecular characterization of melon accessions resistant to Fusarium wilts

Fusarium wilt incited by Fusarium oxysporum f. sp. melonis (F.o.m) is one of the most widespread and devastating melon diseases. While resistance to physiological races 0, 1, and 2 is relatively frequent in different botanical varieties, sources of resistance to race 1,2 are restricted to a few Far-Eastern accessions. In this work, the results of a screening for resistance to F.o.m. race 1,2 among 32 accessions are presented. Three Japanese accessions (‘Kogane Nashi Makuwa’, ‘C-211’, and ‘C-40’) showed the highest resistance levels, but useful levels of resistance were also detected in one Russian ‘C-160’ and two Spanish (‘C-300’ and ‘Mollerusa-7’) accessions. These resistant materials, together with other accessions previously described as resistant to F.o.m. races 0, 1, and/or 2 have been morphologically and molecularly characterized. Based on cluster analysis, these accessions have been grouped according to the botanical subspecies they belong to. Assessment of genetic diversity indicated that the resistant accessions to races 0, 1 and 2, are scattered along the established clusters. On the other hand, high levels of resistance to the race 1,2 could be found only among accessions belonging to Cucumis melo subsp. agrestis, nevertheless, a certain degree of resistance to this race could also be found within some accessions belonging to subsp. melo. As far as we know, this is the first report of resistance to F.o.m race 1,2 found out from the Far-Eastern melon material. Based on fruits characteristics, it appears that several inodurus and cantalupensis accessions could be exploited in breeding programs as resistance sources to F.o.m races 0, 1 and/or 2 for the improvement of these melon types. The accessions with the highest levels of resistance to the race 1,2 appeared to be very distant both molecularly and morphologically from the commercial types. Nevertheless ‘C-160’, ‘C-300’, and ‘Mollerusa-7’ classified as var. inodorus are morphologically very similar to the Spanish commercial types and might be used as resistant sources in breeding these melon types.     

Variance component analysis of plant architectural traits and fruit yield in melon

Summary  A cross was made between a unique highly branched, early flowering line, U. S. Department of Agriculture (USDA) 846-1 (P_1; 7 to 11 lateral branches), and ‘Topmark’ (P₂; 2 to 4 lateral branches), a U.S. Western Shipping melon, to produce an array of 119 F₃ families. Subsequently, a genetic analysis was conducted at Arlington and Hancock, Wisconsin in 2001 to evaluate the segregating progeny for factors likely involved in yield-formation, including days to anthesis, percentage of plants with early pistillate flowering, primary branch number, fruit number and weight per plant, average weight per fruit, percentage of plants with predominantly crown fruit set, and percentage of plants with early maturing fruit. Although, genotype × environment (G × E) interactions were important for some traits (e.g., fruit number and fruit weight), considerable additive and/or dominance variance was detected for all traits. This research provides critical data associated with highly branched melon germplasm including trait correlations and heritabilies (broad- and narrow-sense ranged between 0.28 and 0.91) that used judiciously will allow the development high yielding melon cultivars with early, basally concentrated fruit suitable for once-over or machine harvesting operations.     

Development of molecular markers linked to the <Emphasis Type="Italic">Fom-1</Emphasis> locus for resistance to Fusarium race 2 in melon

Fusarium wilt, caused by Fusarium oxysporum f. sp. melonis (F.o.m), is a worldwide soil-borne disease of melon (Cucumis melo L.). The most effective control measure available is the use of resistant varieties. Resistance to races 0 and 2 of this fungal pathogen is conditioned by the dominant gene Fom-1. An F₂ population derived from the ‘Charentais-Fom1’ × ‘TRG-1551’ cross was used in combination with bulked segregant analysis utilizing the random amplified polymorphic DNA (RAPD) markers, in order to develop molecular markers linked to the locus Fom-1. Four hundred decamer primers were screened to identify three RAPD markers (B17₆₄₉, V01₅₇₈, and V06₁₀₉₂) linked to Fom-1 locus. Fragments amplified by primers B17₆₄₉ and V01₅₇₈ were linked in coupling phase to Fom1, at 3.5 and 4 cM respectively, whereas V06₁₀₉₂ marker was linked in repulsion to the same dominant resistant allele at 15.1 cM from the Fom-1 locus. These RAPDs were cloned and sequenced in order to design primers that would amplify only the target fragment. The derived sequence characterized amplified region (SCAR) markers SB17₆₄₅ and SV01₅₇₄ (645 and 574 bp, respectively) were present only in the resistant parent. The SV06₁₀₉₂ marker amplified a band of 1092 bp only in the susceptible parent. These markers are more universal than the CAPS markers developed by Brotman et al. (Theor Appl Genet 10:337–345, 2005). The analysis of 24 melon accessions, representing several melon types, with these markers revealed that different melon types behaved differently with the developed markers supporting the theory of multiple, independent origins of resistance to races 0 and 2 of F.o.m.     

Genetic analysis of resistance of three melon lines to Sphaerotheca fuliginea

Summary The inheritance of resistance of three melon lines to Sphaerotheca fuliginea race 1 has been studied. Negro possesses one dominant gene for resistance. Resistance in Amarillo is mainly controlled by one gene, and resistance in Moscatel Grande seems to be controlled by two genes. These four genes are different from each other and allelism tests showed that the gene present in Negro is different from those described before in PMR5, PI 124111 and PI 124112. The main gene present in Amarillo is different from those carried by PMR5 and PI 124111 and the two genes carried by Moscatel Grande are different from the one present in PMR5.     

Assessment of genetic variation in a collection of lentil using molecular tools

A collection of lentil landraces, mostly from Italy, but including Mediterranean and foreign germplasm as reference, was evaluated using RAPDs, microsatellite-primed PCR and ISSRs. A low level of useful polymorphic bands was detected with the former two markers, whereas ISSRs revealed a higher degree of variation. The UPGMA trees, based on Jaccard similarity index matrices, obtained with RAPD and ISSR respectively, did not produce similar clusters. However, in both cases, the accession which differentiated the most from the others was from Ethiopia, a country where also other cultivated species have shown a remarkable degree of variation. On the other hand, Italian accessions showed a trend to group together. ISSR markers proved to be useful for distinguishing closely related genotypes, and possibly for substantiating the genetic peculiarity of ecotypes applying for the obtainment of origin and quality marks. This revised version was published online in July 2006 with corrections to the Cover Date.     

A genetic linkage map of Spinacia oleracea and localization of a sex determination locus

A genetic map of Spinach (Spinacia oleracea) was constructed in a classical back cross population using 101 AFLP and 9 microsatellite markers. The map was divided into seven linkage groups with a total length of 585 cM and an average distance between the markers of 5.18 cM. The linkage map was constructed with LOD 3.5, but was quite stable with seven linkage groups remaining until LOD 7.0. Gender segregated 1 male to 1 female in the mapping population and was mapped to a small area of one linkage group with a distance of 1.9 cM to a microsatellite marker termed SO4. This small chromosomal region co-segregating with sex determination in the species is in contrast to previous reports on a heterologous XY chromosome system in spinach. Microsatellite markers used as anchors in the map construction were isolated from sequences of known nuclear encoded genes in spinach. This enabled simultaneous positioning on the map of these genes: Rubisco activase (Rca), Photosytem 1 subunit V (PsaG), Protein Kinase (Pk), Nitrate reductase (Nir), ferrodoxin:thioredoxin reductase (Ftr), Ribosomal protein L1 (Rps22), Choline monooxygenase (Cmo), Pseudogene for BZIP protein (Bzip), Glycerol-3-phosphate acyltransferase (Act1) and stromal ascorbate peroxidase, thylakoid-bound ascorbate peroxidase (Apx2). Spinach has a small genome, which makes it suitable for basic genomic studies and many physiologically important genes have been cloned from the species. The present map anchored with user friendly microsatellite markers will be useful for future studies of physiology and genetics of the species as well as studies of the nature of gender determination.     

Characterization of six varieties of Cucumis melo L. based on morphological and physiological characters, including shelf-life of fruit

Seventy-two accessions covering six varieties of Cucumis melo were characterized by using 35 morphological characters with emphasis on shelf-life, and the relationships between shelf-life and related characters was investigated. Principal component analysis (PCA) revealed that development period of plant and fruit, size of seed and fruit, shelf-life, stem hair, flesh juiciness, netting, abscission of peduncle, rapid yellowing of epidermis at maturity, Brix value, and color of flesh and epidermis etc. were the principal characters to discriminate melon accessions examined in the present study. According to the scatter diagram, vars. acidulusand makuwa, both of which belong to the Oriental melon, are closely related because of their short growth duration, small seed, thin pericarp and poor shelf-life, while American cantaloupe (var.reticulatus) and European cantaloupe (var. cantalupensis) are rather closely related due to their climacteric fruit with orange flesh, slipped peduncle and rapid yellowing of epidermis at maturity, which is closely related with their shelf-life. PCA also indicated that var. saccharinus was closer to var. inodorus than to the other varieties, due to their requirement of long period for development, large size of seed and fruit, and half- or non-slipped peduncle. Shelf-life of melon fruit was significantly correlated with the following characters: quality of flesh, size of seed and fruit, abscission of peduncle, development periods of plant and fruit, rapid yellowing of epidermis at maturity, Brix value and color of flesh and epidermis. Accessions with good shelf-life were mostly found in vars. saccharinus and inodorus. This revised version was published online in July 2006 with corrections to the Cover Date.     

New source of resistance to mosaic virus transmission by Aphis gossypii in melon

The melon accession ‘TGR-1551’ shows a clear and total resistance to the transmission of several isolates of CMV and ZYMV when A. gossypii was used as the vector. The genetic analysis of the progenies obtained crossing this accession with the susceptible Spanish cultivar ‘Bolade Oro’ showed that the resistance to mosaic virus transmission is conferred by the presence of a dominant gene. There are also evidences that this dominant gene is different from ‘Vat’ gene. This revised version was published online in July 2006 with corrections to the Cover Date.     

Climacteric and non-climacteric behavior in melon fruit: 2. Linking climacteric pattern and main postharvest disorders and decay in a set of near-isogenic lines

A set of near-isogenic lines (NILs) of melon (Cucumis melo L.) was used to test the relationship between the climacteric pattern and postharvest disorders at harvest and after 30 days at 8 °C. The NILs contained different chromosome introgressions in the linkage group III from the non-climacteric exotic Korean accession PI 161375 transferred into the genetic background of the non-climacteric Spanish cultivar ‘Piel de Sapo’ (PS). A quantitative trait locus (QTL) in this linkage group induced climacteric behavior in eight NILs accompanied by a peak of ethylene production and fruit dehiscence to different degrees. The cultivar ‘Nicolás’ and one NIL showed a non-climacteric pattern of respiration rate and ethylene production. The climacteric NILs were used to test the relationship between this pattern and postharvest disorders. The reference climacteric lines ‘Fado’ and ‘Védrantais’ were more sensitive to CI and associated Cladosporium rot than the NILs or PS. In general, a more intense climacteric behavior was accompanied by fruit dehiscence, and higher total losses and greater skin scald after storage, than in PS. A higher incidence of chilling injury (CI) in the climacteric NILs was found compared with the non-climacteric ones, although with exceptions (one NIL for CI in the form of scald; the same NIL and one more for pitting). The climacteric onset and netting scald were not related, and CI in the form of skin spots was only found in climacteric NILs and was positively correlated with the maximum peak of ethylene production. Some climacteric NILs did not follow the rule of a higher susceptibility to other disorders and decay after storage compared with PS, such as for example in fruit over-ripening (detected externally or internally), Cladosporium rot at the peduncle and Alternaria rot. Mealiness was independent of climacteric behavior. Three climacteric NILs obtained better flavor scores after storage than PS, although the maximum peak of ethylene production was positively correlated with off-flavor. Genotypic correlation between disorder data and the physiological data of climacteric fruit revealed positive (flavor index) or negative postharvest consequences (skin injuries, rots or off-flavors). At least one QTL can be assigned to most of the quality traits analyzed.     

Inheritance and linkage analysis of resistance to zucchini yellow mosaic virus, watermelon mosaic virus, papaya ringspot virus and powdery mildew in melon

A melon (Cucumis melo L.) breeding line derived from PI 414723 is resistant to three potyviruses,watermelon mosaic virus (WMV), zucchini yellow mosaic virus (ZYMV), papaya ringspot virus (PRSV), and to powdery mildew (PM). The inheritance and linkage relationships of these four resistances were studied in a segregating F2 population and derived F3 families from a cross between cultivar Top Mark and the resistant breeding line. Dominant monogenic inheritance of all four resistances was observed. We report that line 414723-4S3, which was initially selected as a source of ZYMV and WMV resistance, is also a source of dominant monogenic resistances to PRSV and PM race 1. We also report on genetic linkage (significant departure from independent segregation, χ² = 58.1, p≪ 0.0001) between resistance to WMV and ZYMV. The map distance between these loci was estimated to be 7.5 cm. The genes for resistance to PM and PRSV segregated independently from each other, and from ZYMV and WMV resistance. This revised version was published online in July 2006 with corrections to the Cover Date.     

Cultivar identification and genetic relationship among selected breeding lines and cultivars in chickpea (Cicer arietinum L.)

Twenty two RAPD and 22 ISSR markers were evaluated for their potential use in determination of genetic relationships in chickpea (Cicer arietinum L.) cultivars and breeding lines. We were able to identify six chickpea cultivars/breeding lines by cultivar-specific markers. All of the cultivars tested displayed a different phenotype generated either by the RAPD or ISSR primers. Though ISSR primers generated less markers than RAPD primers, the ISSR primers produced higher levels of polymorphism (% of polymorphic markers per primer) than RAPD primers. A high level of within cultivar homogeneity was observed in chickpea. Cultivars/breeding lines originating from a common genetic background showed closer genetic relationship. Chickpea lines with similar seed type(kabuli or desi) had a tendency to cluster together. This revised version was published online in August 2006 with corrections to the Cover Date.     

Screening of melon (Cucumis melo L.) germplasm for multiple disease resistance

Summary Melon germplasm was screened for cucumber green mottle mosaic virus (CGMMV), powdery mildew (Sphaerotheca fuliginea), downy mildew (Pseudoperonospora cubensis) and Fusarium wilt (Fusarium oxysporum f. sp. melonis) resistance under artificial conditions except downy mildew for which screening was done under natural epiphytotic conditions. High level resistance to all the four diseases was not recorded in any of the collections tested. Nevertheless, ertheless, resistance to three diseases was located in three germplasm. Wild Cucumis species C. figarei exhibited absolute resistance to CGMMV and Fusarium wilt and high level resistance to downy mildew. Phoot or snapmelon (Cucumis melo var. momordica) — a non-dessert from of Indian origin—was highly resistant to downy mildew and resistant to CGMMV and medium resistant to Fusarium wilt. Iroquois was resistant to powdery mildew and medium resistant to downy mildew and CGMMV.     

Construction of a genetic linkage map and QTL analysis of fiber-related traits in upland cotton (Gossypium hirsutum L.)

A genetic linkage map with 70 loci (55 SSR, 12 AFLP and 3 morphological loci) was constructed using 117 F₂ plants obtained from a cross between two upland cotton cultivars Yumian 1 and T586, which have relatively high levels of DNA marker polymorphism and differ remarkably in fiber-related traits. The linkage map comprised of 20 linkage groups, covering 525 cM with an average distance of 7.5 cM between two markers, or approximately 11.8% of the recombination length of the cotton genome. The present genetic linkage map was used to identify and map the quantitative trait loci (QTLs) affecting lint percentage and fiber quality traits in 117 F_2:3 family lines. Sixteen QTLs for lint percentage and fiber quality traits were identified in six linkage groups by multiple interval mapping: four QTLs for lint percentage, two QTLs for fiber 2.5% span length, three QTLs for fiber length uniformity, three QTLs for fiber strength, two QTLs for fiber elongation and two QTLs for micronaire reading. The QTL controlling fiber-related traits were mainly additive, and meanwhile including dominant and overdominant. Several QTLs affecting different fiber-related traits were detected within the same chromosome region, suggesting that genes controlling fiber traits may be linked or the result of pleiotropy.     

New molecular markers linked to qualitative and quantitative powdery mildew and scald resistance genes in barley for dry areas

A partial genetic linkage map was constructed on 71 doubled-haploid lines derived from a cross between the barley lines Tadmor and WI2291 with 181 molecular markers. The segregating population was used to detect markers linked to the gene Mlg conferring resistance to powdery mildew (Erysiphe graminis f. sp. hordei) and to genes for quantitative resistance to scald (Rhynchosporium secalis). The gene Mlg on chromosome 4H was flanked by two AFLP markers at a distance of 2.0 and 2.4 cM, respectively. QTLs for resistance to scald were detected on chromosomes 2H and 3H. This association of molecular markers with qualitative and quantitative disease resistance loci represents a valuable starting-point for marker-assisted selection. This revised version was published online in July 2006 with corrections to the Cover Date.     

Assessment of genetic diversity within and among Basmati and non-Basmati rice varieties using AFLP,ISSR and SSR markers

Molecular markers provide novel tools to differentiate between the various grades of Basmati rice, maintain fair-trade practices and to determine its relationship with other rice groups in Oryza sativa. We have evaluated the genetic diversity and patterns of relationships among the 18 rice genotypes representative of the traditional Basmati, cross-bred Basmati and non-Basmati (indica and japonica) rice varieties using AFLP, ISSR and SSR markers. All the three marker systems generated higher levels of polymorphism and could distinguish between all the 18 rice cultivars. The minimum number of assay-units per system needed to distinguish between all the cultivars was one for AFLP, two for ISSR and five for SSR. A total of 171 (110 polymorphic), 240 (188 polymorphic) and 160 (159 polymorphic) bands were detected using five primer combinations of AFLP, 25 UBC ISSR primers and 30 well distributed, mapped SSR markers, respectively. The salient features of AFLP, ISSR and SSR marker data analyzed using clustering algorithms, principal component analysis, Mantel test and AMOVA analysis are as given below: (i) the two traditional Basmati rice varieties were genetically distinct from indica and japonica rice varieties and invariably formed a separate cluster, (ii) the six Basmati varieties developed from various indica × Basmati rice crosses and backcrosses were grouped variably depending upon the marker system employed; CSR30 and Super being more closer to traditional Basmati followed by HKR228, Kasturi, Pusa Basmati 1 and Sabarmati, (iii) AFLP, ISSR and SSR marker data-sets showed moderate levels of positive correlation (Mantel test, r = 0.42–0.50), and (iv) the partitioning of the variance among and within rice groups (traditional Basmati, cross-bred Basmati, indica and japonica) using AMOVA showed greater variation among than within groups using SSR data-set, while reverse was true for both ISSR and AFLP data-sets. The study emphasizes the need for using a combination of different marker systems for a comprehensive genetic analysis of Basmati rice germplasm. The high-level polymorphism generated by SSR, ISSR and AFLP assays described in this study shall provide novel markers to differentiate between traditional Basmati rice supplies from cheaper cross-bred Basmati and long-grain non-Basmati varieties at commercial level.The first two authors have equal contribution     

Potential and limitations of isozymes for chromosomal localization of resistance genes against barley mild mosaic virus (BaMMV)

Summary To assess the possibilities offered by isozymes to locate resistance genes against barley mild mosaic virus (BaMMV), the isozyme patterns of 19 barley (Hordeum vulgare L.) genotypes carrying genes different from ym4 were determined. Of the 15 isozyme systems tested, only three were polymorphic, namely aconitate hydratase, esterase, phosphogluconate dehydrogenase, providing markers on four of the seven barley chromosomes. Studies of F₂ progenies derived from three crosses between resistant genotypes and susceptible varieties failed to reveal linkage between resistance genes and isozymes. Another goal of the experiment was to study the linkage relationships between ym4 and the esterase locus (Est1-Est2-Est4). Our estimates of the recombination rate between these two loci (3.41 and 8.32%) were in the range of those reported between these esterases and one of the resistance genes of the Chinese variety Mokusekko 3.     

Assessment of genetic variability in grapefruits (Citrus paradisi Macf.) and pummelos (C. maxima (Burm.) Merr.) using RAPD and SSR markers

The genetic variability of 38 grapefruit (Citrus paradisi Macf.) and three pummelos (C. maxima (Burm.) Merr..) accessions was evaluated using RAPD, and single sequence repeat (SSR) analyses. Approximately49% of the 198 RAPD were polymorphic, and 4.6 alleles per SSR loci were identified. PIC values changed from 0.093 to 0.450. A UPGMA phenetic tree was constructed and two main grapefruit groups were identified. The grapefruit accessions `do Cabo' and `Siamesa-Filipinas'clustered very close to the pummelos in Group A. The Group B consisted of three sub-groups, which comprised all of the other grapefruit accessions. The majority of grapefruit accessions showed a narrow genetic base suggesting that the observed morphological polymorphism within the group must be associated with somatic mutations, which were not detected by these molecular markers. This revised version was published online in August 2006 with corrections to the Cover Date.     

Localisation and combination of resistance genes against soil-borne viruses of barley (BaMMV,BaYMV) using doubled haploids and molecular markers

Barley yellow mosaic virus disease caused by different strains of BaYMV and BaMMV is a major threat to winter barley cultivation in Europe. Different resistance genes against these viruses have been mapped and suitable PCR-based markers have been developed. In this respect doubled haploid (DH) populations proved to be advantageous as they facilitate a repeated test for resistance against all agents of the barley yellow mosaic virus complex and besides this, dominant marker systems are as informative as co-dominant ones in DHs due to the lack of heterozygous genotypes. Using DH populations resistance genes rym4, rym5, rym11, rym13, rym15 and the BaYMV/BaYMV-2 resistance of the barley cultivar ‘Chikurin Ibaraki 1’ have been mapped. DHs are also well suited to pyramiding resistance genes against BaMMV and BaYMV. Since homozygous recessive genotypes are more frequent in DHs than in segregating F₂ populations, DHs can be efficiently used to create broad-spectrum resistance and to extend the usability of partly overcome resistance genes. Results from employing two different strategies for pyramiding, based on one and two DH-steps, respectively, combining three recessive resistance genes, i.e. rym4/rym5, rym9 and rym11, are presented. The faster strategy based on one haploidy step resulted in the identification of all three and two-way combinations of the respective resistance genes.     

Development of pepper SSR markers from sequence databases

Simple sequence repeat (SSR) markers are potentiallyvaluable tools for plant breeding. The objectives ofthe work reported here were to search the EMBL andGenbank databases for the presence of SSR-containingsequences from the genus Capsicum, to assess thefrequency of different motifs, and to examine thepolymorphism of selected markers in a panel ofgenotypes, including 10 Capsicum spp. and 1tomato and 1 potato genotype. Fifty-eightmicrosatellites with different motifs were found inCapsicum sequences. A subset of twelve of thesewere selected for the polymorphism survey using PCRprimers flanking the SSR. Polymorphisms between Capsicum lines can be detected with 5 of these primerpairs. PCR products of the predicted size were alsoamplified with three primer pairs in potato and oneprimer pair in tomato. The study also showed thatshorter microsatellites could be valuable markers inCapsicum.