ABSTRACT Radish (Raphanus sativus L.) is an important root vegetable in the family Brassicaceae, and is popular worldwide, especially in East Asia. Colchicine was used to induce autotetraploidy in four advanced inbred lines of radish. A combination of 0.1% (w/v) colchicine in 0, 0.1%, 0.2%, or 0.4% (v/v) dimethyl sulphoxide (DMSO) was applied to the apical meristem of young seedlings. A preliminary screening for putative tetraploids was conducted based on morphological traits including flower size, the number of chloroplasts in guard cells, stomatal size, and stomatal density. Plants with significantly larger stomata, with more chloroplasts in their guard cells, were selected for further analysis by chromosome counting. Only one tetraploid genotype, ‘Nau-dy13’, was successfully induced using 0.1% (w/v) colchicine plus 0.2% (v/v) DMSO, with a tetraploid induction rate of 20%. Application of the above concentrations of colchicine and DMSO to the apical meristem of ‘Nau-dy13’ seedlings was found to be an efficient method to induce polyploidy in radish. The tetraploid plants had larger flowers, stomata, and pollen grains and a chromosome number of 4x = 36. In contrast, the rate of pollen germination (19.24%) in the tetraploid genotype was lower than in the diploid line (55.80%). Levels of expression of six genes controlling meiosis, MER3/RCK, ATK1, ATK5, DMC1, TTN8, and MPS1, were measured using real-time reverse transcription quantitative PCR (RT-qPCR). Expression of MPS1 was downregulated 0.6-fold in autotetraploid plants compared to diploid plants. An efficient method for the induction of tetraploids in radish has been established. This will facilitate the manipulation of ploidy level when developing novel elite germplasm and allow further analysis of the mechanism of polyploidy in radish.

Radish (Raphanus sativus L.) is an important worldwide root vegetable crop with high nutrient values and is adversely affected by non-essential heavy metals including chromium (Cr). Little is known about the molecular mechanism underlying Cr stress response in radish. In this study, RNA-Seq technique was employed to identify differentially expressed genes (DEGs) under Cr stress. Based on de novo transcriptome assembly, there were 30,676 unigenes representing 60,881 transcripts isolated from radish root under Cr stress. Differential gene analysis revealed that 2985 uingenes were significantly differentially expressed between Cr-free (CK) and Cr-treated (Cr600) libraries, among which 1424 were up-regulated and 1561 down-regulated. Gene ontology (GO) analysis revealed that these DEGs were mainly involved in primary metabolic process, response to abiotic stimulus, cellular metabolic process and small molecule metabolic process. Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis showed that the DEGs were mainly involved in protein processing in endoplasmic reticulum, starch and sucrose metabolism, amino acid metabolism, glutathione metabolism, drug and xenobiotics by cytochrome P450 metabolism. RT-qPCR analysis showed that the expression patterns of 12 randomly selected DEGs were highly accordant with the results from RNA-seq. Furthermore, many candidate genes including signaling protein kinases, transcription factors and metal transporters, chelate compound biosynthesis and antioxidant system, were involved in defense and detoxification mechanisms of Cr stress response regulatory networks. These results would provide novel insight into molecular mechanism underlying plant responsiveness to Cr stress and facilitate further genetic manipulation on Cr uptake and accumulation in radish.

To elucidate the differences in tolerance and uptake of lead (Pb) among various genotypes in radish (Raphanus sativus L.), eight advanced inbred lines were grown in half-strength Hoagland's nutrient solution with 0, 100, 400, 800 mg L-1 Pb(NO3)(2), respectively. The results showed that the dry weight (DW) of shoot and root were significantly decreased with the increasing of Pb concentrations, indicating that the lead has significant effect on inhibiting growth of different radish genotypes. The lead content in whole plant increased with Pb concentration elevating in the nutrient solution. Among the eight genotypes, lead concentration in shoot varied from 0.13 to 110.68 mg kg(-1), while that in root ranged from 0.67 to 14511.18mg kg(-1). The concentration of lead in root was much higher than that in shoot under various lead treatments, indicating that the lead accumulation in radish was mainly in root. Moreover, significant differences of Pb accumulation were observed among different radish genotypes. The 'Nau-Zdcb05' and 'Nau-Dy05' accumulated more lead than the other genotypes, while relatively less lead was accumulated in 'Nau-Rg05' and 'Nau-Yh05'. In addition, the significantly negative relation was found between root Pb concentration and DW of root and shoot, which is in contrast to the relation between shoot lead concentration and DW of root and shoot.

•Cd responsive DEGs in radish was firstly characterized with transcriptome sequencing.•De novo transcriptome sequencing identified 1496 DEGs under Cd stress in radish.•Expression profiles of Cd-responsive DEGs were validated by RT-qPCR analysis.•Some functional genes including PCS1, MT3 and GSH may be responsible for Cd stress.•A schematic model of Cd-responsive regulatory network in radish was proposed.

In this study, a total of 7 ECA and 4 CAS genes were identiifed in Chinese cabbage by protein se-quence alignment. Phylogenetic analysis of ECA and CAS genes revealed that there were great differences be-tween genes in the each gene family, which suggested that different genes might execute different functions. The expression of ECA and CAS genes was detected in tipburn-susceptive and resistant lines after Ca2+deifcien-cy treatment. The results showed that ECA genes were up-regulated in both lines, and expressed at a higher lev-el in resistant plant than in sensitive plant. However, the expression pattern of different CAS genes was not con-sistent, which might be caused by functional division of different CAS genes. Our study suggested the expression of ECA and CAS was affected by calcium deficiency, and associated with tipburn occurrence in seedling of Chinese cabbage, which provided helpful information for further study the molecular mechanism of tipburn resistance in Chinese cabbage.

[Objectives]To characterize the nitrate reductase gene and reduce the nitrate content,the isolation and expression profiling of nitrate reductase gene(Rs NR) in radish(Raphanus sativus L.) was conducted.[Methods]Based on the unigene sequence from transcriptome database,the c DNA of Rs NR was isolated.Bioinformatics methods were used for sequence analysis.The expression of Rs NR in radish plants treated with different KNO3 concentrations(0,5,20,30 and 50mmol·L-1) and time periods(0,4,8,12 and 24h) were detected using RT-q PCR,and the nitrate content and nitrate reductase activity of radish plants were determined.[Results]The c DNA sequence of Rs NR gene was isolated containing an open reading frame(ORF) of 2 742 bp(Gen Bank accession No.: KM272859).The deduced 913 amino acids sequence showed a 95% identity with NR gene(D38219) in Brassica napus.The maximum Rs NR expression level in leaf and root was induced by 30mmol·L-1NO-3-N.In a short-term induction of 30 mmol·L-1NO-3-N,the expression level of Rs NR reached a maximum at 4h in leaf and root,and then decreased steadily.Moreover,the induction kinetics of NRA,nitrate accumulation and Rs NR expression levels were enhanced in a certain value with increasing nitrate concentration,but decreased later.The NRA,nitrate content and the Rs NR expression level showed a increase trend at 0-4h.In contrast,there was a similar decrease trend of NRA and Rs NR expression levels,and a increase trend of nitrate content at 4-24h.[Conclusions]The results indicate that NO-3 may regulate the expression level of NR gene at the transcriptional level,thus affecting the nitrate content and NRA.

In this study,to explore the relationship between DNA methylation and Pb stress response,the DNA methylation level variations of radish plant in response to Pb stress were investigated. The MSAP ratio in radish seedlings exposed to lead at the concentration of 100,400 and 800 mg·L- 1was 12. 7%,14. 2% and 17. 0%,respectively,which was higher than that in the control. The full methylation levels( CmCGG in double strands) were 9. 7%,10. 2% and12. 8% respectively,while the control was 9. 5%. The level of increasing in MSAP and full methylation indicated that de novo methylation occurred in some 5'-CCGG sites under Pb stress. The total DNA methylation level increased with raising of Pb concentration. Four types of MSAP patterns including de novo methylation,de-methylation,atypical pattern and no changes of methylation pattern were identified among Pb treatments and the control. DNA methylation alteration in radish plants treated with Pb was mainly through de novo methylation. These results provided the theoretical foundation for dissecting the molecular mechanism underlying Pb toxicant and plant response to heavy metal stresses.

MicroRNAs (miRNAs) play vital regulatory roles in plant growth and development. The phase transition from vegetative growth to flowering is crucial in the life cycle of plants. To date, miRNA-mediated flowering regulatory networks remain largely unexplored in radish. In this study, two small RNA libraries from radish leaves at vegetative and reproductive stages were constructed and sequenced by Solexa sequencing. A total of 94 known miRNAs representing 21 conserved and 13 non-conserved miRNA families, and 44 potential novel miRNAs, were identified from the two libraries. In addition, 42 known and 17 novel miRNAs were significantly differentially expressed and identified as bolting-related miRNAs. RT-qPCR analysis revealed that some miRNAs exhibited tissue- or developmental stage-specific expression patterns. Moreover, 154 target transcripts were identified for 50 bolting-related miRNAs, which were predominately involved in plant development, signal transduction and transcriptional regulation. Based on the characterization of bolting-related miRNAs and their target genes, a putative schematic model of miRNA-mediated bolting and flowering regulatory network was proposed. These results could provide insights into bolting and flowering regulatory networks in radish, and facilitate dissecting the molecular mechanisms underlying bolting and flowering time regulation in vegetable crops.

Lead (Pb), a ubiquitous but highly toxic heavy metal (HM), is harmful to human health through various pathways including by ingestion of contaminated vegetables. Radish is a worldwide root vegetable crop with significant health and nutritional benefits. However, little is known about Pb translocation and distribution within radish plants after its uptake by the roots. In this study, Pb stress was induced using Pb(NO3)2 in hydroponic culture, aiming to characterize the transport, ultrastructural localization, and distribution of chemical forms of Pb in different tissues of radish. The results showed that the majority of Pb (85.76-98.72%) was retained in underground organs including lateral roots, root heads and taproot skins, while a small proportion of Pb was absorbed by root flesh (0.44-1.56%) or transported to the shoot (1.28-14.24%). A large proportion of Pb (74.11-99.30%) was integrated with undissolved Pb oxalate, protein and pectates forming Pb-phosphate complexes. Moreover, a low-Pb-accumulating line of radish showed a higher proportion of Pb in water-soluble form compared with a high-Pb-accumulating line. Subcellular distribution analysis showed that a large proportion of Pb was bound to cell wall fraction in lateral roots (71.08-80.40%) and taproot skin (46.22-77.94%), while the leaves and roots had 28.36-39.37% and 27.35-46.51% of Pb stored in the soluble fraction, respectively. Furthermore, transmission electron microscopy (TEM) revealed Pb precipitates in intercellular space, cell wall, plasma lemma and vacuoles. Fractionation results also showed the accumulation of Pb on the cell wall, intercellular space and vacuole, and low uptake of undissolved Pb oxalate, protein, pectates and Pb-phosphate complexes, which might be due to low transport efficiency and Pb tolerance of radish. These findings would provide insight into molecular mechanism of Pb uptake and translocation in radish and facilitate development of low-Pb-content cultivars in root vegetable crops.

BACKGROUND:Radish (Raphanus sativus L.) is an economically important root vegetable crop, and the taproot-thickening process is the most critical period for the final productivity and quality formation. MicroRNAs (miRNAs) are a family of non-coding small RNAs that play an important regulatory function in plant growth and development. However, the characterization of miRNAs and their roles in regulating radish taproot growth and thickening remain largely unexplored. A Solexa high-throughput sequencing technology was used to identify key miRNAs involved in taproot thickening in radish.RESULTS:Three small RNA libraries from 'NAU-YH' taproot collected at pre-cortex splitting stage, cortex splitting stage and expanding stage were constructed. In all, 175 known and 107 potential novel miRNAs were discovered, from which 85 known and 13 novel miRNAs were found to be significantly differentially expressed during taproot thickening. Furthermore, totally 191 target genes were identified for the differentially expressed miRNAs. These target genes were annotated as transcription factors and other functional proteins, which were involved in various biological functions including plant growth and development, metabolism, cell organization and biogenesis, signal sensing and transduction, and plant defense response. RT-qPCR analysis validated miRNA expression patterns for five miRNAs and their corresponding target genes.CONCLUSIONS:The small RNA populations of radish taproot at different thickening stages were firstly identified by Solexa sequencing. Totally 98 differentially expressed miRNAs identified from three taproot libraries might play important regulatory roles in taproot thickening. Their targets encoding transcription factors and other functional proteins including NF-YA2, ILR1, bHLH74, XTH16, CEL41 and EXPA9 were involved in radish taproot thickening. These results could provide new insights into the regulatory roles of miRNAs during the taproot thickening and facilitate genetic improvement of taproot in radish.

Increasing evidence has revealed that microRNA (miRNA)-mediated gene regulation plays a significant role in response to heavy metal stresses. However, there is little information available about the expression patterns or roles of miRNAs under lead toxicity stress in plants. The radish is an important root vegetable crop with a fleshy taproot as the edible part. It was of vital importance to investigate the response mechanisms and explore the regulatory network at the molecular level under the heavy metal stresses in radish. In the present study, using high-throughput sequencing and degradome analysis, a genome-wide identification of radish miRNA and their targets under the exposure of Pb stress was conducted. A total of 74 known and 173 potential novel miRNAs were successfully identified from two radish root libraries of one untreated control (CK) and one Pb-stressed (Pb500). Of these, 25 known and nine novel miRNAs were significantly differentially expressed and identified as Pb-responsive miRNAs. Degradome analysis revealed that 1,979 miRNA-mRNA target transcripts could potentially be cleaved. Gene Ontology (GO) analysis revealed that these target transcripts were predominately involved in the regulation of transcription, defense responses, and binding related terms. The identified target genes for Pb-responsive miRNAs were mainly involved in stress-related signal sensing and transduction, specific metal uptake and homeostasis mechanisms. Additionally, the expression patterns of 20 Pb-responsive miRNAs and six target genes were validated by quantitative real-time PCR (qRT-PCR). These results provide fundamental insights into the miRNA-mediated regulatory networks and molecular mechanisms underlying plant responsiveness to Pb stresses.

为建立萝卜逆转座子间扩增多态性(IRAP)技术体系,基于萝卜Ty1-copia类逆转座子逆转录酶的保守序列设计引物,对IRAP-PCR反应主要因素进行分析.建立的萝卜IRAP标记技术体系(20 μl)为:20 ng基因组DNA模板,1×PCR buffer,0.25 mmol/L dNTPs,2.0 mmol/L Mg2+,0.4 μmol/L引物,1 U Taq DNA聚合酶.将所建立的IRAP标记技术体系应用于14个萝卜品种指纹图谱分析,结果显示,筛选出的2个特异引物RsTy1F5和RsTy1F10在14份萝卜材料中共扩增得到了16个多态性条带,可以将14份萝卜材料完全区分开,每份种质都有独特的指纹图谱,表明IRAP技术可以有效地应用于萝卜种质鉴定和指纹图谱的构建.

BACKGROUND:Salt stress is one of the most representative abiotic stresses that severely affect plant growth and development. MicroRNAs (miRNAs) are well known for their significant involvement in plant responses to abiotic stresses. Although miRNAs implicated in salt stress response have been widely reported in numerous plant species, their regulatory roles in the adaptive response to salt stress in radish (Raphanus sativus L.), an important root vegetable crop worldwide, remain largely unknown.RESULTS:Solexa sequencing of two sRNA libraries from NaCl-free (CK) and NaCl-treated (Na200) radish roots were performed for systematical identification of salt-responsive miRNAs and their expression profiling in radish. Totally, 136 known miRNAs (representing 43 miRNA families) and 68 potential novel miRNAs (belonging to 51 miRNA families) were identified. Of these miRNAs, 49 known and 22 novel miRNAs were differentially expressed under salt stress. Target prediction and annotation indicated that these miRNAs exerted a role by regulating specific stress-responsive genes, such as squamosa promoter binding-like proteins (SPLs), auxin response factors (ARFs), nuclear transcription factor Y (NF-Y) and superoxide dismutase [Cu-Zn] (CSD1). Further functional analysis suggested that these target genes were mainly implicated in signal perception and transduction, regulation of ion homeostasis, basic metabolic processes, secondary stress responses, as well as modulation of attenuated plant growth and development under salt stress. Additionally, the expression patterns of ten miRNAs and five corresponding target genes were validated by reverse-transcription quantitative PCR (RT-qPCR).CONCLUSIONS:With the sRNA sequencing, salt-responsive miRNAs and their target genes in radish were comprehensively identified. The results provide novel insight into complex miRNA-mediated regulatory network of salt stress response in radish, and facilitate further dissection of molecular mechanism underlying plant adaptive response to salt stress in root vegetable crops.

The fu11-1ength coding sequence of f1avono1 synthase gene (DaFLS1) in purp1e yam was obtained by using 3’-and 5’-RACE methods. DaFLS1 ( GenBank acc. no:KJ022640 ) , is 1 113 bp in 1ength, containing an open reading frame of 1 005 bp encoding 334 amino acids. The DaFLS1 protein is water so1ub1e without signa1 peptide and transmembrane domain, 1oca1ized in cytop1asm. Conserved domain search revea1ed DaFLS1 protein be1onged to the 2OG-Fe(II) oxygenase superfami1y ,whose activity depended on the co-existence ofα-ketog1utaric acid and ferrousion. Drtho1ogy of the deduced amino acid sequence showed DaFLS1 was high1y homo1ogous with Narcissus tazetta ( AFS63900,75% simi-1arity), and Allium cepa(AAT68476,74% identity). The phy1ogenetic tree constructed based on the deduced amino acid sequence indicated yam had far re1ationships with oth-er p1ants. DaFLS1 exhibited the highest expression 1eve1 in young 1eaf with two peaks over the who1e growth period. There was 1itt1e or no expression in other tissues. The expression of DaFLS1 gene in purp1e yam shared the same features of FLSs expression in other p1ants.

利用40个EST-SSR标记,对100份甘蓝自交系组成的自然群体进行遗传结构分析,采用TASSEL3.0软件的GLM (general linear model)和MLM (mixed linear model)模型,对甘蓝中心柱长和中心柱长/球高比值性状进行关联分析.结果显示供试材料可分为3个亚群;EST-SSR位点间有较高的多态性和一定程度的连锁不平衡;以GLM模型分析,共检测出5个标记的7个位点与中心柱长性状相关联,2个标记的3个位点与中心柱长/球高性状相关联,其中2个位点同时与两个性状相关联;以MLM模型分析,共检测出3个标记的4个位点与中心柱长性状相关联,1个标记的2个位点与中心柱长/球高性状相关联,其中2个位点同时与两个性状相关联.GLM和MLM两种模型同时检测到了与中心柱长/球高相关联的1个标记的2个位点以及与中心柱长相关联的3个标记的4个位点.

microRNAs (miRNAs) are a class of single-stranded endogenous non-coding RNAs that play critical roles in plant growth, development, and environmental stress responses. Temperature is one of the major physical parameters disturbing cellular homeostasis and causing leaf etiolation in plants. Previous studies have reported that several conserved and novel miRNAs were responsive to heat stress in plants. However, the characterization of miRNAs responsive to heat stress in radish remains poorly understood. To better understand miRNAs and their target genes under heat stress, two small RNA libraries were constructed from heat-treated (Heat24) and heat-untreated (CK) radish roots. Using Solexa system, totally, 26 known and 19 novel miRNAs were identified as differentially expressed under heat stress. Expression patterns of a set of heat-responsive miRNAs were validated by quantitative real-time PCR (qRT-PCR). Furthermore, 422 sliced targets for 25 known miRNAs were identified by degradome sequencing technology, and most of the identified targets are involved in multiple biological processes including transcriptional regulation and response to biotic and abiotic stresses. Moreover, some miRNAs and their corresponding targets, which are related to the accumulation of heat stress transcription factors and heat shock proteins, played important roles in thermo-tolerance in radish. These findings could enhance the understanding of molecular mechanisms underlying miRNAs and their targets in regulating plant responses to heat stress.

采用电子克隆与基因组步移策略分离出萝卜RsFPF1基因gDNA和cDNA及启动子序列,并进行表达特征分析及转基因功能验证.序列分析表明,RsFPF1基因长度为330 bp,编码109个氨基酸;蛋白同源分析表明,RsFPF1蛋白与拟南芥及白芥FPF1蛋白间亲缘关系最近.RsFPF1基因5 '上游启动子区序列长度为1 845 bp,采用PLACE和PlantCARE软件分析表明,该启动子序列含有典型调控元件及多个光响应顺式元件.半定量RT-PCR表达分析表明,开花前RsFPF1基因在茎尖表达量最高,开花后在花及花蕾中表达量最高.通过农杆菌介导的遗传转化获得转RsFPF1基因的烟草阳性植株,与野生型相比,转入RsFPF1基因的植株出现花期提前现象.结论:RsFPF1基因能够促进萝卜提早开花,其表达可能受光调控,在调控萝卜抽薹开花及花发生相关基因表达方面发挥着重要作用.

Simple sequence repeats (SSRs) are highly polymorphic and co-dominant markers, providing an important genomic resource for genetic research. Recently, large-scale transcriptome sequencing has become a reliable and efficient approach for the identification and development of new genic-SSR markers and has been successfully conducted in a few important plant species. However, SSR development based on transcriptome sequencing remains limited in radish (Raphanus sativus L.). In the present study, from a total of 73,084 unigenes and 150,455 contigs which were assembled from 71.95 million Illumina sequence reads of a radish taproot library, a collection of 11,928 genic-SSR loci were successfully identified in 11,311 unigene sequences. Trinucleotide repeats were the most abundant repeat units, as in many other plants, with a frequency of 52 %. Furthermore, a total of 5,503 genic-SSR primers were developed, from which 1,052 SSR primers were synthesized, and a subset of 823 (78.23 %) primers could generate stable bands. Moreover, 67 selected informative genic-SSR markers were used to determine the genetic diversity of 32 radish genotypes, in which the polymorphism information content values ranged from 0.49 to 0.89. For effective cultivar identification, a novel strategy called manual cultivar identification diagram was employed. Thirty-two radish accessions were clearly separated by six genic-SSR markers. Additionally, the cross-species/genera transferability of these SSRs was further validated in nine relatives in Brassicaceae. These results suggested that the novel genic-SSR markers, as a basis for future genetic linkage and gene tagging analysis, could be very valuable in facilitating genetic mapping, marker-assisted selection and comparative genome analysis.

Myrosinase is a defense-related enzyme and is capable of hydrolyzing glucosinolates into a variety of compounds, some of which are toxic to pathogens and herbivores. Many studies revealed that a number of important vegetables or oil crops contain the myrosinase-glucosinolate system. However, the related promoter and genomic DNA sequences as well as expression profiles of myrosinase gene remain largely unexplored in radish (Raphanus sativus). In this study, the 2 798 bp genomic DNA sequence, designated as RsMyr2, was isolated and analyzed in radish. The RsMyr2 consisting of 12 exons and 11 introns reflected the common gene structure of myrosinases. Using the genomic DNA walking approach, the 5′-flanking region upstream of RsMyr2 with length of 1 711 bp was successfully isolated. PLACE and PlantCARE analyses revealed that this upstream region could be the promoter of RsMyr2, which contained several basic cis-regulatory elements including TATA-box, CAAT-box and regulatory motifs responsive to defense and stresses. Furthermore, recombinant pET-RsMyr2 protein separated by SDS-PAGE was identified as myrosinase with mass spectrometry. Real-time PCR analysis showed differential expression profiles of RsMyr2 in leaf, stem and root at different developmental stages (e.g., higher expression in leaf at cotyledon stage and lower in flesh root at mature stage). Additionally, the RsMyr2 gene exhibited up-regulated expression when treated with abscisic acid (ABA), methyl jasmonate (MeJA) and hydrogen peroxide (H2O2), whereas it was down-regulated by wounding (WO) treatment. The findings indicated that the expression of RsMyr2 gene was differentially regulated by these stress treatments. These results could provide new insight into elucidating the molecular characterization and biological function of myrosinase in radish.

Downy mildew (DM), caused by the fungus Peronospora parasitica, is a destructive disease of radish (Raphanus sativus L.) worldwide. Host resistance has been considered as an attractive and environmentally friendly approach to control the disease. However, the genetic mechanisms of resistance in radish to the pathogen remain unknown. To determine the inheritance of resistance to DM, F1, F2 and BC1F1 populations derived from reciprocal crosses between a resistant line NAU-dhp08 and a susceptible line NAU-qtbjq-06 were evaluated for their responses to DM at seedling stage. All F1 hybrid plants showed high resistance to DM and maternal effect was not detected. The segregation for resistant to susceptible individuals statistically fitted a 3:1 ratio in two F2 populations (F2(SR) and F2(RS)), and 1:1 ratio in two BC1F1 populations, indicating that resistance to DM at seedling stage in radish was controlled by a single dominant locus designated as RsDmR. A total of 1972 primer pairs (1036 SRAP, 628 RAPD, 126 RGA, 110 EST-SSR and 72 ISSR) were screened, and 36 were polymorphic between the resistant and susceptible bulks, and consequently used for genotyping individuals in the F2 population. Three markers (Em9/ga24370, NAUISSR826700 and Me7/em10400) linked to the RsDmR locus within a 10.0 cM distance were identified using bulked segregant analysis (BSA). The SRAP marker Em9/ga24370 was the most tightly linked one with a distance of 2.3 cM to RsDmR. These markers tightly linked to the RsDmR locus would facilitate marker-assisted selection and resistance gene pyramiding in radish breeding programs.