Construction Of Genetic Linkage Map Research Articles

Generation and analysis of blueberry transcriptome sequences from leaves, developing fruit, and flower buds from cold acclimation through deacclimation

BackgroundThere has been increased consumption of blueberries in recent years fueled in part because of their many recognized health benefits. Blueberry fruit is very high in anthocyanins, which have been linked to improved night vision, prevention of macular degeneration, anti-cancer activity, and reduced risk of heart disease. Very few genomic resources have been available for blueberry, however. Further development of genomic resources like expressed sequence tags (ESTs), molecular markers, and genetic linkage maps could lead to more rapid genetic improvement. Marker-assisted selection could be used to combine traits for climatic adaptation with fruit and nutritional quality traits.ResultsEfforts to sequence the transcriptome of the commercial highbush blueberry (Vaccinium corymbosum) cultivar Bluecrop and use the sequences to identify genes associated with cold acclimation and fruit development and develop SSR markers for mapping studies are presented here. Transcriptome sequences were generated from blueberry fruit at different stages of development, flower buds at different stages of cold acclimation, and leaves by next-generation Roche 454 sequencing. Over 600,000 reads were assembled into approximately 15,000 contigs and 124,000 singletons. The assembled sequences were annotated and functionally mapped to Gene Ontology (GO) terms. Frequency of the most abundant sequences in each of the libraries was compared across all libraries to identify genes that are potentially differentially expressed during cold acclimation and fruit development. Real-time PCR was performed to confirm their differential expression patterns. Overall, 14 out of 17 of the genes examined had differential expression patterns similar to what was predicted from their reads alone. The assembled sequences were also mined for SSRs. From these sequences, 15,886 blueberry EST-SSR loci were identified. Primers were designed from 7,705 of the SSR-containing sequences with adequate flanking sequence. One hundred primer pairs were tested for amplification and polymorphism among parents of two blueberry populations currently being used for genetic linkage map construction. The tetraploid mapping population was based on a cross between the highbush cultivars Draper and Jewel (V. darrowii is also in the background of 'Jewel'). The diploid mapping population was based on a cross between an F1 hybrid of V. darrowii and diploid V. corymbosum and another diploid V. corymbosum. The overall amplification rate of the SSR primers was 68% and the polymorphism rate was 43%.ConclusionsThese results indicate that this large collection of 454 ESTs will be a valuable resource for identifying genes that are potentially differentially expressed and play important roles in flower bud development, cold acclimation, chilling unit accumulation, and fruit development in blueberry and related species. In addition, the ESTs have already proved useful for the development of SSR and EST-PCR markers, and are currently being used for construction of genetic linkage maps in blueberry.

Diversity Array Technology Markers: Genetic Diversity Analyses and Linkage Map Construction in Rapeseed (Brassica napus L.)

We developed Diversity Array Technology (DArT) markers for application in genetic studies of Brassica napus and other Brassica species with A or C genomes. Genomic representation from 107 diverse genotypes of B. napus L. var. oleifera (rapeseed, AACC genomes) and B. rapa (AA genome) was used to develop a DArT array comprising 11 520 clones generated using PstI/BanII and PstI/BstN1 complexity reduction methods. In total, 1547 polymorphic DArT markers of high technical quality were identified and used to assess molecular diversity among 89 accessions of B. napus, B. rapa, B. juncea, and B. carinata collected from different parts of the world. Hierarchical cluster and principal component analyses based on genetic distance matrices identified distinct populations clustering mainly according to their origin/pedigrees. DArT markers were also mapped in a new doubled haploid population comprising 131 lines from a cross between spring rapeseed lines ‘Lynx-037DH’ and ‘Monty-028DH’. Linkage groups were assigned on the basis of previously mapped simple sequence repeat (SSRs), intron polymorphism (IP), and gene-based markers. The map consisted of 437 DArT, 135 SSR, 6 IP, and 6 gene-based markers and spanned 2288 cM. Our results demonstrate that DArT markers are suitable for genetic diversity analysis and linkage map construction in rapeseed.

Two EST-derived marker systems for cultivar identification in tree peony

Tree peony (Paeonia suffruticosa Andrews), a woody deciduous shrub, belongs to the section Moutan DC. in the genus of Paeonia of the Paeoniaceae family. To increase the efficiency of breeding, two EST-derived marker systems were developed based on a tree peony expressed sequence tag (EST) database. Using target region amplification polymorphism (TRAP), 19 of 39 primer pairs showed good amplification for 56 accessions with amplicons ranging from 120 to 3,000bp long, among which 99.3% were polymorphic. In contrast, 7 of 21 primer pairs demonstrated adequate amplification with clear bands for simple sequence repeats (SSRs) developed from ESTs, and a total of 33 alleles were found in 56 accessions. The similarity matrices generated by TRAP and EST-SSR markers were compared, and the Mantel test (r=0.57778, P=0.0020) showed a moderate correlation between the two types of molecular markers. TRAP markers were suitable for DNA fingerprinting and EST-SSR markers were more appropriate for discriminating synonyms (the same cultivars with different names due to limited information exchanged among different geographic areas). The two sets of EST-derived markers will be used further for genetic linkage map construction and quantitative trait locus detection in tree peony.

Novel genomic and EST-derived SSR markers in Japanese chestnuts

A total of 366 novel simple sequence repeat (SSR) markers were developed in Japanese chestnut ( Castanea crenata), comprising 220 genomic SSRs derived from enriched genomic libraries and 146 expressed sequence tag (EST)–SSRs obtained from large-scale EST sequencing analysis. Thirty accessions, comprising Japanese, Chinese ( C. mollissima), European ( Castanea sativa), and American chestnuts ( Castanea dentata), were used for evaluation of SSR polymorphism and transferability across species. The EST–SSRs showed less polymorphism than the genomic SSRs and were more transferable. The mean observed heterozygosity ( H O) and the mean expected heterozygosity ( H E) of genomic SSRs in the Japanese chestnuts were 0.63 and 0.68, respectively; those of EST-SSRs were each 0.47. Although about 80% of the genomic SSRs were amplified in all 4 species, more than 95% of the EST–SSRs were transferable across all 4 species. The many novel SSRs developed in this study will be applicable for the construction of genetic linkage maps, QTL analysis of phenotypic traits, high-throughput genotyping of marker-assisted selection, and association genetics.

English

  In this study, we reported 36 novel polymorphic microsatellite markers derived from a cDNA library of Scylla paramamosain. The polymorphisms of these markers were detected in a sample of 24 individuals. A total of 159 alleles were detected with an average of 4.4 alleles per locus. The number of alleles per locus ranged from two to eight. The observed and expected heterozygosity per locus ranged from 0.100 to 1.000 and from 0.097 to 0.819, with an average of 0.679 and 0.625, respectively. Five loci significantly deviated from Hardy-Weinberg equilibrium after Bonferroni correction (P < 0.0014) and no significant linkage disequilibrium between pairs of loci was found. These loci will provide useful information for the studies on genetic diversity and structure, construction of genetic linkage maps and the effectively management of S. paramamosain.   Key words: Scylla paramamosain, microsatellite markers, cDNA, polymorphism.

Analysis of adaptive responses of Pinus pinaster to changing environmental conditions in the Mediterranean region

Analysis of adaptive responses of Pinus pinaster to changing environmental conditions in the Mediterranean region

Mapping and transcriptomic approches implemented for understanding disease resistance to Phytophthora cinammomi in Castanea sp

The European chestnut, Castanea sativa Mill, covers a total area of 2.53 million hectares two million of which are chestnut forests, i.e. forests where chestnut is the dominant tree species, being the remaining 0.53 million hectares devoted to fruit production (20.9% of the total chestnut-growing area). Chestnut fruit production in Europe declined considerably during the XX century to the current 200,000 t (almost 300 million euros). This decline arose mainly due to serious diseases and changes in the structure of society. Towards the end of the century, there was a sharp increase in chestnut demand which triggered new planting and the restoration of old orchards throughout Europe, although ink disease (Phytophthora spp.), canker blight (Cryphonectria parasitica (Murrill) M.E. Barr.) and more recently Dryocosmus kuriphilus Yasumatsu, still represent major threats to the species. In Europe, chestnut breeding has been focused on the improvement of cultivars and rootstocks through selection and hybridisation with Asian species resistant to ink disease. Considerable success was obtained using this approach, however little information was acquired until now on the genetic basis of resistance in chestnut. A set of interspecific crosses were established between European chestnut and Asian species and two separate full-sib pedigrees have been produced, SC (C. sativa x C. crenata) and SM (C.sativa x C. mollissima). The goal is to perform DNA marker:trait association analysis to identify QTLs related to ink disease metrics and also to identify putative resistance genes to P cinnamomi using a transcriptomic approach. An overview of the study will be presented including the genotyping of the mapping population (parents and progenies) with nuclear SSR markers designed for Castanea mollissima and provided by the American team of Fagaceae Genomics Project, for the construction of the first genetic linkage map. The methodologies implemented for the determination of the metrics of resistance of mapping population, for further identification of QTLs, will also be presented.

Genetic dissection of transcript, metabolite, growth and wood property traits in an F2 pseudo-backcross pedigree of Eucalyptus grandis x E. urophylla

Background E. grandis is used extensively for the production of pulp and paper due to its rapid growth, good form and ease of vegetative propagation. E. urophylla exhibits tolerance to fungal diseases that limit the growth of E. grandis in tropical and subtropical regions. Interspecific hybrids of these two species are, therefore, commonly used to produce fast-growing, disease tolerant hybrids for clonal eucalypt plantations in tropical and subtropical regions (e.g., South Africa, Congo and Brazil). These hybrids often exhibit superior growth and wood quality compared to the pure species, but the underlying genetic basis of the observed hybrid superiority remains unclear. Phenotypic variation observed in interspecific mapping populations has been used to identify QTLs in several genetic linkage studies in Eucalyptus. However, QTL intervals have generally been wide (20 to 30 cM) and may include several hundred genes. To bridge the gap between fine mapping and QTL validation studies, the expression levels of genes and metabolites in individuals from the segregating population can be treated as quantitative traits and used for eQTL and mQTL mapping, respectively. Co-localization of wood property, expression and metabolite QTLs will facilitate the identification of positional candidate genes and other components of regulatory networks underlying phenotypic variation. Methods To identify genetic factors controlling growth and wood property traits in Eucalyptus, an F2 pseudo-backcross mapping family derived from a cross between an F1 hybrid (GUSAP1, E. grandisxE. urophylla, Sappi Forest Research) and an E. urophylla parent (USAP1), consisting of 555 individuals, was used for genetic linkage map construction using microsatellite and DArT markers. Phenotypic trait assessment included physical measurements of tree diameter and wood density performed on 319 three-year-old individuals. Klason (acid-soluble & -insoluble) lignin and cell wall sugar content were determined for a selection of 100 backcross progeny and used for near-infrared analysis (NIRA) calibration. NIRA predictions for glucose, xylose, arabinose, cellulose and total lignin content, as well as pulp yield were made for all 315 individuals. Total lignin and S:G ratios were also separately measured for the 315 individuals. Immature xylem tissues, collected from 192 backcross progeny, were used for metabolite profiling (ORNL, Oak Ridge, TN) and Illumina mRNA-Seq (PE50, BGI Americas) quantification of transcript levels.

High-density genetic linkage maps with over 2,400 sequence-anchored DArT markers for genetic dissection in an F2 pseudo-backcross of Eucalyptus grandis × E. urophylla

Traits that differentiate cross-fertile plant species can be dissected by genetic linkage analysis in interspecific hybrids. Such studies have been greatly facilitated in Eucalyptus tree species by the recent development of Diversity Arrays Technology (DArT) markers. DArT is an affordable, high-throughput marker technology for the construction of high-density genetic linkage maps. Eucalyptus grandis and Eucalyptus urophylla are commonly used to produce fast-growing, disease tolerant hybrids for clonal eucalypt plantations in tropical and subtropical regions. We analysed 7,680 DArT markers in an F2 pseudo-backcross mapping pedigree based on an F1 hybrid clone of E. grandis and E. urophylla. A total of 2,440 markers (31.7%) were polymorphic and could be placed in linkage maps of the F1 hybrid and two pure-species backcross parents. An integrated genetic linkage map was constructed for the pedigree resulting in 11 linkage groups (n = 11) with 2,290 high-confidence (LOD ≥ 3.0) markers and a total map length of 1,107.6 cM. DNA sequence analysis of the mapped DArT marker fragments revealed that 43% were located in protein coding regions and 90% could be placed in the recently completed draft genome assembly of E. grandis. Together with the anchored genomic sequence information, this linkage map will allow detailed genetic dissection of quantitative traits and hybrid fitness characters segregating in the F2 progeny and will facilitate the development of markers for molecular breeding in Eucalyptus.

Isolation and characterization of microsatellite markers in the grass Poa arachnifera Torr

Microsatellite markers represent a proven class of genetic markers that have wide use in a variety of plant genetic studies and exhibit several advantages when compared to other DNA-based marker systems. Because of their successful use for genetic analysis in a number of plant species, their generation and availability in the genus Poa (Poaceae) would be advantageous for a multitude of Poa improvement, selection or marker-assisted breeding endeavors. However, due to the cost of their development, few microsatellite markers are available for Poa. In this study, we describe the isolation and characterization of 46 informative microsatellite primer pairs from an unenriched Poa arachnifera Torrey (Texas bluegrass) genomic library. Most of the microsatellite primer pairs amplify a single amplification product and were found to be useful across seven other Poa species. The availability of these informative microsatellite markers should serve a valuable role in the analysis and construction of genetic linkage maps and future marker assisted selection projects.

Genetic linkage map construction for kenaf using SRAP, ISSR and RAPD markers

With 3 figures and 3 tablesAbstractKenaf (Hibiscus cannabinus L.) is one of the most economically important crops for fibre production. Genetic maps have been developed in almost all the agricultural crops. However, genetic information for kenaf, especially, at the molecular level is limited. To better understand the genetic basis of kenaf for the improvement of production and to lay the foundation for molecular breeding efforts, a primary genetic linkage map was constructed using sequence‐related amplified polymorphism (SRAP), inter‐simple sequence repeat (ISSR) and randomly amplified polymorphic DNA (RAPD). Cultivar ‘Alian kenaf’ and ‘Fuhong 992’ were used as parents to construct an F2 population consisting of 180 plants. We selected 494 SRAP, 60 ISSR, 120 RAPD and 300 two‐primer RAPD mixture primers that amplified 396 polymorphic loci in total. At a logarithm of the odds (LOD) score threshold of 5.0 and at a maximum map distance of 25 cM, these 396 loci were used to construct the genetic linkage map with MAPMAKER/EXP 3.0, a total of 307 loci were grouped into 26 linkage groups that spanned a total map length of around 4924.8 cM with a mean density of 16.04 cM per locus. These markers were distributed randomly in all linkage groups without any clustering. The construction of the kenaf genetic linkage map will be useful for further genetic studies including mapping both qualitative and quantitative traits, marker‐assisted selection program, and comparative genomics analysis.

A Note on Development of a Low-cost and High-throughput SSR-based Genotyping Method in Peanut (Arachis hypogaea L.)

ABSTRACT Peanut cultivar development has been dominated by conventional breeding methods, which have and will continue to play an important role. Applications of marker-assisted selection (MAS) have been used in peanut breeding selection but the cost of genotyping is still a considerable factor. The objective of this study was to introduce a simple, low-cost, and high-throughput protocol for peanut community. The developed system was based on a smaller (10.5 cm in length) polyacrylamide gel size system to separate PCR amplified DNA fragments and silver staining to visualize the bands. This system is very easy to operate, having one electrophoresis unit holds two vertical 52-sample gels, and the cost for purchasing the unit is less than $200. For instance, the electrophoresis runs about 1 hr and 40 min at 180 V for 9% polyacrylamide gel to separate small to medium sized DNA bands (< 500 bp) or 1 hr and 20 min at 160 V for 6% polyacrylamide gel preferably for larger band separation (≥ 500 bp), but the gel concentrations and running times could be adjusted according to the polymorphic banding patterns and sizes to mitigate the drawback of this system of small gel-size. The silver staining takes about 30 min. After staining, the gels are placed on a light-box for genotype scoring and then photographed using a digital camera. The cost per gel is estimated at $0.54 and the cost for silver staining is estimated at $0.37. Therefore, the cost could be as low as $0.018 per data point, excluding PCR reaction and DNA preparation cost. A scientist has the potential to generate over 1,200 data points per day. This method has been used in the construction of a peanut genetic linkage map and QTL studies in our laboratory in conjunction with other methods.

Isolation and characterization of 16 polymorphic microsatellite markers from Nibea albiflora

Nibea albiflora is a commercially important fish species in China. Herein we report 16 novel polymorphic microsatellite markers in Nibea albiflora by using the 5′ anchored polymerase chain reaction (PCR) technique. The characteristics of these loci were estimated by using a sample of 30 individuals. A total of 79 alleles were detected with an average of 4.9 alleles per locus. The number of alleles per locus ranged from three to nine. The polymorphism information content (PIC) values for the 16 microsatellite loci ranged from 0.3131 to 0.7910. The observed and expected heterozygosity per locus ranged from 0.2333 to 1.000 and from 0.3452 to 0.8421, with an average of 0.7248 and 0.6592, respectively. Four loci significantly deviated from Hardy–Weinberg equilibrium after Bonferroni correction (P < 0.0031), and no significant linkage disequilibrium between pairs of loci was found. This study provides useful information for studies on genetic diversity and structure, construction of genetic linkage maps of N. albiflora, and effective management of this fish resource.

Identification of type I microsatellite markers associated with genes and ESTs in Scylla paramamosain

In this study, Microsatellites were screened from genes and ESTs of Scylla paramamosain and the related species Scylla serrata. Eighteen polymorphic loci were detected, with a total of 106 alleles. The observed and expected heterozygosity per locus ranged from 0.08 to 1.00, and from 0.08 to 0.88. Polymorphism information content (PIC) showed that 12 loci were high informative (PIC > 0.5). Two loci were found to deviate significantly from Hardy–Weinberg equilibrium after Bonferroni correction (P < 0.0028). Significant linkage disequilibrium was detected between loci Scpa-ALF-SSR and Scpa-SSR03 after Bonferroni correction (P < 0.0003). Cross-species amplification of these 18 microsatellite markers showed that most of them could give the expected products in three other species: S. serrata, Scylla tranquebarica and Scylla olivacea. This study will be useful for construction of genetic linkage maps, mapping of quantitative trait loci (QTL), and analysis of comparative genome in S. paramamosain and the related taxa.

Development of 57 novel polymorphic microsatellite markers in half-smooth tongue sole (Cynoglossus semilaevis)

Half-smooth tongue sole (Cynoglossus semilaevis) is a promising species for aquaculture in China. The wild population of C. semilaevis is under threat from environmental factors. Microsatellite markers are very suitable for assessing genetic diversity. Four microsatellite-enriched libraries of half smooth tongue sole (Cynoglossus semilaevis) were constructed, from which 57 polymorphic microsatellites were isolated and characterized. The polymorphism of these microsatellites was assessed by genotyping in 30 individual fish. The number of alleles ranged from 2 to 11, with an average of 4.614 alleles per locus. The values of observed and expected heterozygosities ranged from 0.1000 to 1.0000 and from 0.0966 to 0.8847 respectively. Polymorphism information content (PIC) ranged from 0.0905 to 0.862. These markers would be useful for population structure assessment, genetic linkage map construction and parentage analysis for this species.

A sequence-based genetic linkage map as a reference for Brassica rapa pseudochromosome assembly

BackgroundBrassica rapa is an economically important crop and a model plant for studies concerning polyploidization and the evolution of extreme morphology. The multinational B. rapa Genome Sequencing Project (BrGSP) was launched in 2003. In 2008, next generation sequencing technology was used to sequence the B. rapa genome. Several maps concerning B. rapa pseudochromosome assembly have been published but their coverage of the genome is incomplete, anchoring approximately 73.6% of the scaffolds on to chromosomes. Therefore, a new genetic map to aid pseudochromosome assembly is required.ResultsThis study concerns the construction of a reference genetic linkage map for Brassica rapa, forming the backbone for anchoring sequence scaffolds of the B. rapa genome resulting from recent sequencing efforts. One hundred and nineteen doubled haploid (DH) lines derived from microspore cultures of an F1 cross between a Chinese cabbage (B. rapa ssp. pekinensis) DH line (Z16) and a rapid cycling inbred line (L144) were used to construct the linkage map. PCR-based insertion/deletion (InDel) markers were developed by re-sequencing the two parental lines. The map comprises a total of 507 markers including 415 InDels and 92 SSRs. Alignment and orientation using SSR markers in common with existing B. rapa linkage maps allowed ten linkage groups to be identified, designated A01-A10. The total length of the linkage map was 1234.2 cM, with an average distance of 2.43 cM between adjacent marker loci. The lengths of linkage groups ranged from 71.5 cM to 188.5 cM for A08 and A09, respectively. Using the developed linkage map, 152 scaffolds were anchored on to the chromosomes, encompassing more than 82.9% of the B. rapa genome. Taken together with the previously available linkage maps, 183 scaffolds were anchored on to the chromosomes and the total coverage of the genome was 88.9%.ConclusionsThe development of this linkage map is vital for the integration of genome sequences and genetic information, and provides a useful resource for the international Brassica research community.

English

A mapping population comprising 220 F2&nbsp;individuals was developed by raising the selfed F1&nbsp;seed of a single hybrid plant, White Ponni x Moroberekan. Two hundred and twenty two F3&nbsp;families were phenotyped for blast resistance under artificial condition. Fifty random amplified polymorphic (RAPD) primers were used to identify parental polymorphism. Twelve RAPD markers which showed clear polymorphism were scored in the F2&nbsp;mapping population. Out of 12 markers, 11 loci (91.97 %) fitted into the expected segregation ratio of 3:1 based on&nbsp;c2&nbsp;-&nbsp;test at 0.05% probability value. Four hundred microsatellite primer pairs were used in the parental survey, Out of which 52 simple sequence repeats (SSR) primers were polymorphic. Out of 52 marker loci, 44 marker loci (84.61%) fitted into the expected Mendelian segregation ratio of 1:2:1 based on&nbsp;c2&nbsp;-&nbsp;test at 0.05% probability value. The multipoint analysis resulted in the construction of preliminary genetic linkage map with 40 loci. One way analysis of variance (ANOVA) was performed to identify the marker phenotype association which resulted in the identification of 23 SSR markers putatively associated for the six traits studied. Three SSR markers (RM 5757, RM 451 and RM 492 from chromosomes four and two) were linked for leaf blast resistance, six markers for days to flowering, nine markers for pigmentation trait, two markers for plant height, four markers for panicle length and seven markers linked for grain yield per plant. Out of 12 RAPD markers studied, 7 were found to be linked to three traits viz., leaf blast resistance (OPBB 5258,&nbsp;OPBB 5194, OPAL 16940&nbsp;and OPBD 12680), five markers for glume purple tip (GPT) and one marker for grain yield per plant based on single marker analysis. &nbsp; Key words:&nbsp;Magnaporthe grisea&nbsp;(Hebert) Barr,&nbsp;Oryza sativa. L., marker assisted selection (MAS).

A first set of gene-derived polymorphic microsatellite markers in half-smooth tongue sole (Cynoglossus semilaevis)

Half-smooth tongue sole (Cynoglossus semilaevis) is a commercially important marine fish species in China. A set of type I microsatellite markers were identified through bioinformatic mining of the GenBank database. Thirteen of these markers showed polymorphisms through genotyping a sample of 30 individuals. A total of 47 alleles were detected, with an average of 3.6 alleles per locus. The number of alleles, observed and expected heterozygosity per locus ranged from two to five, from 0.14 to 0.93 and from 0.27 to 0.77, respectively. Three loci significantly deviated from Hardy-Weinberg equilibrium after Bonferroni correction (P < 0.0038) and no significant linkage disequilibrum between pairs of loci was found. The markers identified in this study will contribute to construction of genetic linkage maps and mapping of quantitative trait loci (QTL) of C. semilaevis.

Characterization and genetic linkage mapping of the horticulturally important mutation leafless inflorescence ( lli) in periwinkle Catharanthus roseus

Catharanthus roseus is a seasonal to perennial garden plant and the exclusive source of the anticancer drugs vincristine and vinblastine. Its horticultural importance is due to the compound racemose inflorescence architecture of branches in which pairs of prominent flowers are subtended by one of the two leaves per node. Here is reported the construction of a molecular framework genetic linkage map and mapping on it of the LEAF-LESS INFLORESCENCE ( LLI) locus. It is quantitatively shown that the adult lli mutant plants produce altered inflorescence of improved horticultural value, wherein axes are excessively branched, two flowers are formed per node that are bare of subtending leaves, and several times more open flowers are displayed each day, as compared to LLI plants. Segregational analysis of 914 PCR-based polymorphic markers identified 564 of them that segregated according to monogenic 1:1 ratio among 191 F 2:7 recombinant inbred lines of a mutant mapping population. The linkage analysis of 564 DNA markers gave a map in which 84 RAPD, 11 ISSR and 39 SSR (anonymous) markers and 8 microRNA and 30 EST-SSR (coding region) markers were organized into 8 linkage groups (LGs). All LGs contained 2–20 microsatellite markers. The framework map spanned 1790.4 cM and intermarker distance was 10.3 cM. While known primers/primer pairs were used to develop RAPD, ISSR, SSR and microRNA markers, the EST database of the species was mined for EST-SSR markers. EST-SSR resources developed in the study have added to the existing repertoire of microsatellites in C. roseus. The lli locus mapped to a 3.9 cM interval in the middle of LG4 flanked by microsatellite markers.

Application of molecular marker techniques in rye research

Rye(Secale cereal L.) is an important alien gene resources in wheat(Triticum aestivum L.) genetic research as it is a related species of Triticinae.It has been used in improving disease resistance,yield and grain quality of wheat.Molecular marker technique(a valuable tool in molecular biological research) has been used extensively in rye research.This review summarized the progress in research regarding the applications of molecular marker techniques in rye.Such techniques included genetic linkage map construction,valuable gene identification and mapping,and specific rye genome marker development and application.The application prospects of molecular marker techniques in rye research were also discussed.