Development of sequence-characterized amplified regions (SCARs) from amplified fragment length polymorphism (AFLP) markers tightly linked to the Vf gene in apple.

Amplified fragment length polymorphism (AFLP) markers have become widely used in saturating the region of a gene of interest for the ultimate goal of map-based cloning of the gene or for marker-assisted selection. However, conversion of AFLP markers into restriction fragment length polymorphism (RFLP) or polymerase chain reaction (PCR)-based markers will greatly expand their usefulness in genetic applications. Previously, we have identified 15 AFLP markers tightly linked to the Vf gene conferring scab resistance in apple. In this study, we have successfully converted 11 of these AFLPs into sequence-characterized amplified region (SCAR) markers. Of the remaining four nonconverted AFLP markers, one, ET2MC8-1, has been found to be very short (83 base pairs) and is an A/T rich (90%) marker; a second, EA2MG11-1, has shown identical sequences between Malus floribunda 821 (the original source of the Vf gene) and scab-susceptible apple cultivars; while the other two, EA12MG16-1 and ET8MG1-1, have not been cloned. Using the 11 converted SCAR markers along with 5 previously identified SCAR markers, a high-resolution linkage map around the Vf gene has been constructed, and found to be consistent with its corresponding AFLP map. Three converted SCAR markers (ACS-3, -7, and -9) are inseparable from the Vf gene; whereas one (ACS-6) is located left of, and the remaining seven (ACS-1, -2, -4, -5, -8, -10, and -11) are located right of the Vf gene at genetic distances of 0.4 and 0.2 cM, respectively. A reliable and robust procedure for development of SCAR markers from AFLP markers is presented.     

AFLP-derived SCARs facilitate construction of a 1.1 Mb sequence-ready map of a region that spans the Vf locus in the apple genome

The availability of high-density anchored markers is a prerequisite for reliable construction of a deep coverage BAC contig, which leads to creation of a sequence-ready map in the target chromosomal region. Unfortunately, such markers are not available for most plant species, including woody perennial plants. Here, we report on an efficient approach to build a megabase-size sequence-ready map in the apple genome for the Vf region containing apple scab resistance gene(s) by targeting AFLP-derived SCAR markers to this specific genomic region. A total of 11 AFLP-derived SCAR markers, previously tagged to the Vf locus, along with three other Vf-linked SCAR markers have been used to screen two apple genome BAC libraries. A single BAC contig which spans the Vf region at a physical distance of approximately 1,100 kb has been constructed by assembling the recovered BAC clones, followed by closure of inter-contig gaps. The contig is 4 ×deep, and provides a minimal tiling path of 16 contiguous and overlapping BAC clones, thus generating a sequence-ready map. Within the Vf region, duplication events have occurred frequently, and the Vf locus is restricted to the ca. 290 kb region covered by a minimum of three overlapping BAC clones.     

Saturation mapping of the apple scab resistance gene Vf using AFLP markers

Using the amplified fragment length polymorphism (AFLP) technique combined with a ”narrow-down” bulk segregant strategy enabled us to quickly identify 15 tightly linked AFLP markers to the Vf gene that confers resistance to the apple scab disease. High-resolution mapping placed all 15 AFLP markers within an interval of 0.6 cM around the Vf region; 7 of them were found to be inseparable from the Vf gene, 1 was localized left of, and the remaining 7 located right of the Vf gene. In addition, eight previously identified RAPD markers were also mapped, but only three, including M18, AM19, and AL07, were localized within this short interval, and none co-segregated with the Vf gene. The saturation of the Vf region with AFLP markers will accelerate both marker-assisted selection and map-based cloning. The advantages of this ”narrow-down” strategy, estimation of physical distances among AFLP markers, and their potential application are also discussed. Received: 22 December 1999 / Accepted: 25 March 2000     

Phenotypic Reaction and Genetic Analysis Using AFLP-derived SCARs for Resistance to Apple Scab

Six sequence‐characterized amplified region (SCAR) markers linked to the apple scab resistance gene Vf were evaluated for their utility in marker‐assisted selection (MAS) in apple breeding. Of the six SCARs used in this study, ACS‐6 was located left of the Vf gene, ACS‐7 and ACS‐9 co‐segregated with Vf, and ACS‐8, ACS‐4, ACS‐5 were located right of the Vf gene. Three families derived from crosses between scab‐resistant and scab‐susceptible cultivars, including ‘Liberty’ × ‘Deljub’, ‘Liberty’ × ‘Delcorf’, and ‘Florina’ בDelcorf’, previously screened for scab resistance following greenhouse inoculation with the fungal pathogen Venturia inaequalis, were genotyped and compared with phenotypic reactions to scab infection in the field. For each family, a subset progeny of 30 seedlings (propagated onto Malling 9 rootstock and of 7 years old) was selected based on fungal sporulation according to the following scheme. Ten seedlings with no visible scab sporulation on leaves were given phenotypic scores of 0 (deemed resistant); 10 seedlings with moderate scab sporulation were given phenotypic scores of 1.0 (deemed moderately resistant); and 10 seedlings with heavy sporulation were given phenotypic scores of 2.0 (deemed susceptible). DNA was isolated from leaf tissue collected from all 90 seedlings, parents and Malus floribunda 821, the original source of the Vf gene, and screened with all six SCARs. All six SCARs were present in the two scab‐resistant parents, ‘Liberty’ and ‘Florina’, and M. floribunda 821; while, the two scab‐susceptible parents, ‘Deljub’ and ‘Delcorf’, lacked all SCARs. All SCARs were either present or absent in varying numbers of seedlings in each progeny with phenotypic ratings of either 0 (resistant) or 1.0 (moderately resistant); while all seedlings with phenotypic ratings of 2.0 (susceptible) lacked all SCARs. The inconsistencies between phenotypic scab ratings and SCAR marker data are discussed.     

Development of an STS map of an interspecific progeny of Malus

Simple sequence repeat (SSR) markers developed from Malus, as well as Prunus, Pyrus and Sorbus, and some other sequence-tagged site (STS) loci were analysed in an interspecific F₁ apple progeny from the cross ‘Fiesta’ × ‘Totem’ that segregated for several agronomic characters. A linkage map was constructed using 259 STS loci (247 SSRs, four SCARs and eight known-function genes) and five genes for agronomic traits—scab resistance (Vf), mildew resistance (Pl-2), columnar growth habit (Co), red tissues (Rt) and green flesh background colour (Gfc). Ninety SSR loci and three genes (ETR1, Rt and Gfc) were mapped for the first time in apple. The transferability of markers from other Maloideae to Malus was found to be around 44%. The loci are spread across 17 linkage groups, corresponding to the basic chromosome number of Malus and cover 1,208 cM, approximately 85% of the estimated length of the apple genome. Interestingly, we have extended the top of LG15 with eight markers covering 25 cM. The average map density is 4.7 cM per marker; however, marker density varies greatly between linkage groups, from 2.5 in LG14 to 8.9 in LG7, with some areas of the genome still in need of further STS markers for saturation. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. An erratum to this article can be found at     

Towards the map-based cloning of Vf: fine and physical mapping of the Vf Region

A map-based cloning scheme is being used to isolate the Vf resistance gene of apple against the fungus Venturia inaequalis. Vf is a major dominant gene that is inherited in a Mendelian manner and influenced by minor genes that modify its activity. The two recently published markers M18 and AL07, bracketing Vf, were tested on 1179 progeny plants of three crosses to fine-map Vf. M18 and AL07 were positioned at 0.2 cM and 1.1 cM from Vf respectively, for a total distance between the two markers of 1.3 cM. Physical mapping by pulsed-field gel electrophoresis, using M18 and AL07 as probes, demonstrated that both markers hybridize to a common 870 kb NotI restriction fragment. We therefore found a relationship between physical and genetic distance of 670 kb/cM in the Vf region. This led us to the conclusion that a chromosome walk using the recently published apple BAC library starting from M18 and AL07 is feasible. Received: 8 February 1999 / Accepted: 16 March 1999     

Multiple field and glasshouse assessments increase the reliability of linkage mapping of the Vf source of scab resistance in apple

 Apple scab, caused by the fungus Venturia inaequalis (Cke.) Wint., is an important disease in commercial apple production. A mapping population of 155 individuals, derived from a cross between the apple varieties ‘Prima’ (resistant)בFiesta’ (susceptible), was scored for response to the disease in replicated field and glasshouse trials throughout Europe. Twenty data sets were selected and cluster analysis was used to form a consensus score for the population fitting a 1 : 1 segregation ratio of resistance:susceptibility. The progeny were scored with molecular markers. A detailed map covering 54 cM of the ‘Prima’ linkage group containing the Vf gene for scab resistance was constructed using 24 molecular markers linked to the resistance gene. One isoenzyme marker (Pgm-1), six RFLP markers and 17 RAPD markers formed a linkage group with the consensus measure of resistance to scab. Four marker bridges were established with the corresponding ‘Fiesta’ linkage group with additional markers (one isozyme, one RFLP, three RAPD and one AFLP). A low chi-square value indicated a good fit of the marker ordering, which was in close agreement with previously reported linkage positions for some of the markers and Vf. Differences were observed in the ability of different scoring methods to resolve susceptible and resistant classes. The results obtained for the consensus classification of resistance to scab for the population may suggest the presence of virulent inocula at some sites, which could overcome the Vf gene for resistance. The consequences of relying on individual scoring occasions for studying Vf scab resistance are discussed in the context of linkage analysis, conventional breeding selection, and marker-assisted selection. Received: 23 July 1997 / Accepted: 31 October 1997     

Inheritance and molecular mapping of two fertility-restoring loci for Honglian gametophytic cytoplasmic male sterility in rice (<Emphasis Type="Italic">Oryza sativa </Emphasis>L.)

The Honglian cytoplasmic male sterility (cms-HL) system, a novel type of gametophytic CMS in indica rice, is being used for the large-scale commercial production of hybrid rice in China. However, the genetic basis of fertility restoration (Rf) in cms-HL remains unknown. Previous studies have shown that fertility restoration is controlled by a single locus located on chromosome 10, close to the loci Rf1 and Rf4, which respond to cms-BT and cms-WA, respectively. To determine if the Rf locus for cms-HL is different from these Rf loci and to establish fine-scale genetic and physical maps for map-based cloning of the Rf gene, high-resolution mapping of the Rf gene was carried out using RAPD and microsatellite markers in three BCF₁ populations. The results of the genetic linkage analysis indicated that two Rf loci respond to cms-HL, and that these are located in different regions of chromosome 10. One of these loci, Rf5 , co-segregates with the SSR marker RM3150, and is flanked by RM1108 and RM5373, which are 0.9 cM and 1.3 cM away, respectively. Another Rf locus, designated as Rf6(t), co-segregates with RM5373, and is flanked by RM6737 and SBD07 at genetic distances of 0.4 cM. The results also demonstrated these loci are distinct from Rf1 and Rf4. A 105-kb BAC clone covering the Rf6(t) locus was obtained from a rice BAC library. The sequence of a 66-kb segment spanning the Rf6(t) locus was determined by a BLASTX search in the genomic sequence database established for the cultivar 93-11.Communicated by R. Hagemann     

Molecular tagging and genetic mapping of the disease resistance gene<Emphasis Type="Italic"> RppQ</Emphasis> to southern corn rust

Southern corn rust (SCR), Puccinia polysora Underw, is a destructive disease in maize (Zea mays L.). Inbred line Qi319 is highly resistant to SCR. Results from the inoculation test and genetic analysis of SCR in five F₂ populations and five BC₁F₁ populations derived from resistant parent Qi319 clearly indicate that the resistance to SCR in Qi319 is controlled by a single dominant resistant gene, which was named RppQ. Simple sequence repeat (SSR) analysis was carried out in an F₂ population derived from the cross Qi319×340. Twenty SSR primer pairs evenly distributed on chromosome10 were screened at first. Out of them, two primer pairs, phi118 and phi 041, showed linkage with SCR resistance. Based on this result, eight new SSR primer pairs surrounding the region of primers phi118 and phi 041 were selected and further tested regarding their linkage relation with RppQ. Results indicated that SSR markers umc1,318 and umc 2,018 were linked to RppQ with a genetic distance of 4.76 and 14.59 cM, respectively. On the other side of RppQ, beyond SSR markers phi 041 and phi118, another SSR marker umc1,293 was linked to RppQ with a genetic distance of 3.78 cM. Because the five linkage SSR markers (phi118, phi 041, umc1,318, umc 2,018 and umc1,293) are all located on chromosome 10, the RppQ gene should also be located on chromosome 10. In order to fine map the RppQ gene, AFLP (amplified fragment length polymorphism) analysis was carried out. A total 54 AFLP primer combinations were analyzed; one AFLP marker, AF1, from the amplification products of primer combination E-AGC/M-CAA, showed linkage with the RppQ gene in a genetic distance of 3.34 cM. Finally the RppQ gene was mapped on the short arm of chromosome 10 between SSR markers phi 041 and AFLP marker AF1 with a genetic distance of 2.45 and 3.34 cM respectively.Communicated by H. F. Linskens     

Screening apples for OPD20/600 using sequence-specific primers

Apple scab, caused by Venturia inaequalis (Cke.) Wint., is the most serious disease of apple trees in many areas of the world. Resistance to V. inaequalis, derived from the small-fruited species Malus floribunda 821, is determined by a major dominant gene, Vf. Using random decamer primers, we identified a RAPD marker, OPD20/600, which is linked to the Vf gene. OPD20/600 was then cloned and sequenced. Sequence-specific primers based on the marker were used to further screen M. floribunda 821, 7 scab-susceptible apple cultivars, 10 scab-resistant apple cultivars, and 28 scab-resistant Coop selections. The sequence-specific primers allowed identification of polymorphisms of OPD20/600 based on the presence or absence of a single band. The advantages of sequence-specific primers over decamer primers for developing genetic markers are discussed.     

苹果Co基因的SSR标记定位（简报）

Columnar apple is an important genetic resource for tree form breeding of apple. In this study, 106 individuals of the F1 population derived from 'Spur Fuji' (coco)x 'Telamon'(Coco) were used as plant materials for screening SSR markers linked to gene. By bulked segregating analysis (BSA), eight SSR markers from the tenth linkage group of apple genome were tested. Finally, three of them, COL, CH02a10 and CH03d11, were identified to be SSR markers of Co gene. Linkage analysis showed that the genetic distance of COL, CH02a10 and CH03d11 to Co locus was 15.3cM, 22.2cM and 3.9 cM, respectively. On the linkage map of these markers, Co gene was located between COL and CH03d11.     

Construction of genetic linkage map of the medicinal and ornamental plant <Emphasis Type="Italic">Catharanthus roseus</Emphasis>

An integrated genetic linkage map of the medicinal and ornamental plant Catharanthus roseus, based on different types of molecular and morphological markers was constructed, using a F₂ population of 144 plants. The map defines 14 linkage groups (LGs) and consists of 131 marker loci, including 125 molecular DNA markers (76 RAPD, 3 RAPD combinations; 7 ISSR; 2 EST-SSR from Medicago truncatula and 37 other PCR based DNA markers), selected from a total of 472 primers or primer pairs, and six morphological markers (stem pigmentation, leaf lamina pigmentation and shape, leaf petiole and pod size, and petal colour). The total map length is 1131.9 cM (centiMorgans), giving an average map length and distance between two markers equal to 80.9 cM and 8.6 cM, respectively. The morphological markers/genes were found linked with nearest molecular or morphological markers at distances varying from 0.7 to 11.4 cM. Linkage was observed between the morphological markers concerned with lamina shape and petiole size of leaf on LG1 and leaf, stem and petiole pigmentation and pod size on LG8. This is the first genetic linkage map of C. roseus.     

RAPD markers linked to the Vf gene for scab resistance in apple

Scab (Venturia inaequalis) is one of the most harmful diseases of apple, significantly affecting world apple production. The identification and early selection of resistant genotypes by molecular markers would greatly improve breeding strategies. Bulked segregant analysis was chosen for the identification of RAPD markers linked to the Vf scab resistant gene. Five different RAPD markers, derived from the wild species Malus floribunda. 821, were identified, and their genetic distance from Vf gene was estimated. The markers OPAM19₂₂₀₀ and OPAL07₅₈₀ were found to be very closely linked to the Vf gene. This result was indirectly confirmed by the analysis of resistant genotypes collected from various breeding programmes. Except for cv Murray, which carries the Vm gene, all these resistant genotypes showed the markers OPAM19₂₂₀₀ and OPAL07₅₈₀.     

A genetic linkage map of <Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> L. and localization of genes for specific resistance to six races of anthracnose (<Emphasis Type="Italic">Colletotrichum lindemuthianum</Emphasis>)

A genetic map of common bean was constructed using 197 markers including 152 RAPDs, 32 RFLPs, 12 SCARs, and 1 morphological marker. The map was established by using a F₂ population of 85 individuals from the cross between a line derived from the Spanish landrace Andecha (Andean origin) and the Mesoamerican genotype A252. The resulting map covers about 1,401.9 cM, with an average marker distance of 7.1 cM and includes molecular markers linked to disease resistance genes for anthracnose, bean common mosaic virus, bean golden yellow mosaic virus, common bacterial blight, and rust. Resistance to races 6, 31, 38, 39, 65, and 357 of the pathogenic fungus Colletotrichum lindemuthianum (anthracnose) was evaluated in F₃ families derived from the corresponding F₂ individuals. The intermediate resistance to race 65 proceeding from Andecha can be explained by a single dominant gene located on linkage group B1, corresponding to the Co-1 gene. The recombination between the resistance specificities proceeding from A252 agrees with the assumption that total resistance to races 6, 31, 38, 39, 65, and 357, is organized in two clusters. One cluster, located on B4 linkage group, includes individual genes for specific resistance to races 6, 38, 39, and 357. The second cluster is located on linkage group B11 and includes individual genes for specific resistance to races 6, 31, 38, 39, and 65. These two clusters correspond to genes Co-3/Co-9 and Co-2, respectively. It is concluded that most anthracnose resistance Co- genes, previously described as single major genes conferring resistance to several races, could be organized as clusters of different genes conferring race-specific resistance. C. Rodríguez-Suárez and B. Méndez-Vigo equally share for authorship.     

A high-resolution map of the H1 locus harbouring resistance to the potato cyst nematode Globodera rostochiensis

The resistance gene H1 confers resistance to the potato cyst nematode Globodera rostochiensis and is located at the distal end of the long arm of chromosome V of potato. For marker enrichment of the H1 locus, a bulked segregant analysis (BSA) was carried out using 704 AFLP primer combinations. A second source of markers tightly linked to H1 is the ultra-high-density (UHD) genetic map of the potato cross SH × RH. This map has been produced with 387 AFLP primer combinations and consists of 10,365 AFLP markers in 1,118 bins (). Comparing these two methods revealed that BSA resulted in one marker/cM and the UHD map in four markers/cM in the H1 interval. Subsequently, a high-resolution genetic map of the H1 locus has been developed using a segregating F₁ SH × RH population consisting of 1,209 genotypes. Two PCR-based markers were designed at either side of the H1 gene to screen the 1,209 genotypes for recombination events. In the high-resolution genetic map, two of the four co-segregating AFLP markers could be separated from the H1 gene. Marker EM1 is located at a distance of 0.2 cM, and marker EM14 is located at a distance of 0.8 cM. The other two co-segregating markers CM1 (in coupling) and EM15 (in repulsion) could not be separated from the H1 gene.Communicated by J.G. Wenzel     

Identification and mapping of the novel apple scab resistance gene Vd3

Apple scab, caused by the fungal pathogen Venturia inaequalis, is one of the most devastating diseases for the apple growing in temperate zones with humid springs and summers. Breeding programs around the world have been able to identify several sources of resistance, the Vf from Malus floribunda 821 being the most frequently used. The appearance of two new races of V. inaequalis (races 6 and 7) in several European countries that are able to overcome the resistance of the Vf gene put in evidence the necessity of the combination of different resistance genes in the same genotype (pyramiding). Here, we report the identification and mapping of a new apple scab resistance gene (Vd3) from the resistant selection “1980-015-25” of the apple breeding program at Plant Research International, The Netherlands. This selection contains also the Vf gene and the novel V25 gene for apple scab resistance. We mapped Vd3 on linkage group 1, 1 cM to the south of Vf in repulsion phase to it. Based on pedigree analysis and resistance tests, it could be deduced that 1980-015-25 had inherited Vd3 from the founder “D3.” This gene provides resistance to the highly virulent EU-NL-24 strain of race 7 of V. inaequalis capable of overcoming the resistance from Vf and Vg. JMS and SGJ contributed equally to this work     

大豆重组自交系群体（ZKS-HX）遗传图谱的构建和形态标记的定位

以大豆品种‘合丰25’为母本,半野生大豆‘新民6号’为父本杂交得到的F2-9代122个重组自交系为试验材料,构建了含有124个SSR标记、1个EST标记、3个形态学标记的大豆遗传图谱。此图谱覆盖的基因组长度为2348．3cM．标记间平均距离为18．3cM。每个连锁群长度范围为15．1～195．9cM之间,标记数范围2—10个。本文将控制茸毛色（Pb）基因定位于LG06-C2连锁群上,与Sat_40x2的遗传距离为39．6cM;控制叶耳g（Le）、花色（4W,）基因定位于LG12-F连锁群上,它们之间的遗传距离为9．9cM,与两边的Satt348、Sat_240标记遗传距离分别为13．3cM和10．5cM。     

Genetic mapping of AFLP/AMF-derived DNA markers in the vicinity of the self-incompatibility locus in<Emphasis Type="Italic"> Ipomoea trifida</Emphasis>

Sporophytic self-incompatibility of diploid Ipomoea trifida is controlled by a single multiallelic locus, the S-locus. To make a fine linkage map around the S-locus, AFLP (amplified restriction fragment length polymorphism) and AMF (AFLP-based mRNA fingerprinting) analyses were performed using bulked genomic DNA and mRNA, respectively, from several plants of each S-haplotype in a segregating population. Putative S-haplotype-specific fragments were obtained and subjected to RFLP analysis of genomic DNA to confirm genetic linkage to the S-locus. Eight DNA markers co-segregating with the S-haplotype were identified and mapped in close proximity to the S-locus. One of them, AAM-68, was the most tightly linked to the S-locus, because no recombinants were detected in the 873 plants of the segregating population analyzed. The S-locus region was defined to be within 1.25 cM in the linkage map. These markers are useful for positional cloning of the S-locus genes in Ipomoea.     

A genetic map of Xenopus tropicalis

We present a genetic map for Xenopus tropicalis, consisting of 2886 Simple Sequence Length Polymorphism (SSLP) markers. Using a bioinformatics-based strategy, we identified unique SSLPs within the X. tropicalis genome. Scaffolds from X. tropicalis genome assembly 2.0 (JGI) were scanned for Simple Sequence Repeats (SSRs); unique SSRs were then tested for amplification and polymorphisms using DNA from inbred Nigerian and Ivory Coast individuals. Thus identified, the SSLPs were genotyped against a mapping cross panel of DNA samples from 190 F2 individuals. Nearly 4000 SSLPs were genotyped, yielding a 2886-marker genetic map consisting of 10 major linkage groups between 73 and 132 cM in length, and 4 smaller linkage groups between 7 and 40 cM. The total effective size of the map is 1658 cM, and the average intermarker distance for each linkage group ranged from 0.27 to 0.75 cM. Fluorescence In Situ Hybridization (FISH) was carried out using probes for genes located on mapped scaffolds to assign linkage groups to chromosomes. Comparisons of this map with the X. tropicalis genome Assembly 4.1 (JGI) indicate that the map provides representation of a minimum of 66% of the X. tropicalis genome, incorporating 758 of the approximately 1300 scaffolds over 100,000 bp. The genetic map and SSLP marker database constitute an essential resource for genetic and genomic analyses in X. tropicalis.     

Genetic mapping of the pear scab resistance gene Vnk of Japanese pear cultivar Kinchaku

Pear scab (caused by Venturia nashicola) is one of the most harmful diseases of pears, especially Japanese and Chinese pear species. The molecular identification and early selection of resistant plants could greatly improve pear breeding. We have identified the position of the scab resistance gene, designated Vnk in an indigenous Japanese pear cultivar Kinchaku, within the pear genome by using simple sequence repeat (SSR) markers derived from pear and apple. The position of Vnk was identified in the central region of linkage group 1 of Kinchaku. Several amplified fragment length polymorphism (AFLP) markers linked to Vnk were obtained by bulked segregant analysis. Among them, the AFLP marker closest to Vnk was converted into a sequence tagged site (STS) marker. Four random amplified polymorphic DNA (RAPD) markers previously found to be loosely associated with Vnk (Iketani et al. 2001) were successfully converted into STS markers. Six markers (one SSR Hi02c07 and five STSs converted from AFLP and RAPD) showed tight linkages to Vnk, being mapped with distances ranging from 2.4 to 12.4 cM. The SSR CH-Vf2, which was isolated from a BAC clone of the contig containing the apple scab gene Vf, was mapped at the bottom of linkage group 1 in Kinchaku, suggesting that the Vnk and Vf loci are located in different genomic regions of the same homologous linkage group.