Polymorphism and DNA markers for asparagus cultivars identified by random amplified polymorphic DNA

Summary DNA polymorphism among five Asparagus officinalis L. cultivars-Imperial, Snow, Steline, UC-157 and Larac, as detected by random amplified polymorphic DNA (RAPD), is reported. Thirty one decamer primers were tested. and twenty six of them yielded amplification products. Fourteen primers gave products with at least one polymorphic DNA fragment. Among a total of 119 amplified fragments 33 were polymorphic. These RAPD markers enabled the identification of asparagus cultivars. Unique markers for cultivars were: Snow-bands 475 bp, 772 bp, 412 bp, 935 bp and 820 bp amplified by primers D5, OPA-07, OPA-09, OPA-10 and OPA-18, respectively. Steline-bands 645 bp, 680 bp and 997 bp amplified by primers A32, OPA-03 and OPA-09, respectively. A band 903 bp, amplitied by primer OPA-12, is a marker for Imperial, and a band 420 bp, amplified by primer D52, is a marker for Larac. Cultivar UC-157 could be identified by a combination of shared polymorphic bands. The pairwise marker difference between cultivars ranged from 0.08 to 0.17. A phenogram of the genetic relationship based on RAPD fits with the known origin of the cultivars.     

Random amplified polymorphic DNA variation among German and Dutch asparagus cultivars

DNA polymorphism among nine cultivars of Asparagus officinalis L. was measured using random amplified polymorphic DNA (RAPD). Of 69 reproducible amplification products from 12 arbitrary decamer primers, 49 RAPD markers were polymorphic and could be used to distinguish six German and three Dutch asparagus cultivars. Even with very small sample sizes, genetic similarity measurements based on the RAPD data allowed accurate grouping of the nine cultivars into distinct clusters, with the exception of two individuals which clustered to closely related varieties. Two German cultivars showed high genetic similarity and were distinct from the remaining German varieties. The German and Dutch cultivars were clearly separated by a relatively large genetic distance.     

Evaluation of random amplified polymorphic DNA (RAPD) markers in Hevea brasiliensis

The applicability of random amplified polymorphic DNA (RAPD) markers in the cultivated rubber tree, Hevea, was evaluated using 43 decamer oligonucleotide primers in a set of 24 clones selected in different South-East Asian countries. A total of 220 0.35–3.5 kb DNA fragments were amplified, of which 111 were polymorphic. Of these, 80 fragments (RAPD markers) which were repeatable and clearly scorable across all genotypes were used to estimate genetic distances among the clones tested. The estimated genetic distances ranged from 0.05 (RRII 308 and PB 5/51) to 0.75 (RRIC 100 and SCATC 88–13). A mean genetic distance of 0.5 indicates a rather high genetic variability among the tested clones. As expected, because of the breeding history of Hevea, UPGMA cluster analysis and Principal Coordinate Analysis (PCoA) indicated the absence of a distinct geographical grouping. The possible application of RAPD markers for clone identification and also for analysis of genetic relationships among Hevea clones is discussed.     

Mapping of Molecular Markers on Brassica B-Genome Chromosomes Added to Brassica napus

Using interspecific hybridization among various Brassica species, B-genome chromosomes from different sources of Brassica, i.e. B. nigra (BB, 2n = 18), B. carinata (BBCC), 2n = 34) and B. juncea (AABB, 2n = 36) were transferred into the Canadian variety ‘Andor’ of B. napus. Monosomic addition lines were selected (AACC + 1B, 2n = 39) by cytological control. For characterization of the alien chromosomes, series of isozymes, RFLPs and RAPD markers were employed. This permitted the identification of a total of 39 lines representing seven of the eight B-genome chromosomes.     

Genetic diversity and identification of cymbidium cultivars as measured by random amplified polymorphic DNA (RAPD) markers

DNA from thirty-six cymbidium cultivars was examined using polymerase chain reaction (PCR) to determine the efficiency of randomly amplified polymorphic DNA (RAPD) markers in identifying cultivars and determining levels of genetic variability. A total of 132 RAPD markers, 78% of which were polymorphic, were produced from 15 10mer arbitrary primers. All the cultivars were distinguishable when a number of primers was considered. One cultivar, Blue Smoke ‘Green Meadow’ could be distinguished from all the rest based only on lack of the OPA5-370 fragment. Genetic distances among the cultivars were estimated based on the amount of band sharing and ranged from 0.08–0.50 with an average of 0.29. Cluster analysis of genetic distance estimates grouped siblings together with each other and parents with offsprings, thereby agreeing with known parentage information and corroborating isozyme data obtained from a separate study. The possible application of the observed polymorphism and variation to cymbidium breeding is discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Identification of commercial chicory cultivars for hydroponic forcing and their phenetic relationships revealed by random amplified polymorphic DNAs and amplified fragment length polymorphisms

One‐hundred and twenty‐four amplified fragment length polymorphism (AFLP) and 49 random amplified polymorphic DNA (RAPD) markers have been used to distinguish between 20 and 23 commercial chicory cultivars, respectively. These were all Cichorium intybus var. foliosum F₁ hybrids, currently used in hydroponic forcing. Five‐hundred and twenty RAPD primers (OPERON) were tested, of which 156 resulted in reproducible patterns and 26 yielded polymorphisms. Two‐hundred and fifty‐six AFLP primer‐combinations were tested and six combinations were selected for identification purposes. Similarity indices were measured and clustering has been done using pairwise comparison. Both types of marker provide similar conclusions. Two major clusters are formed, representing late and early cultivars. All cultivars were identified using 10 informative RAPD primers or three AFLP primer combinations. A low degree of polymorphism was detected between some early cultivars, suggesting a narrow genetic base in their breeding strategy.     

Genetic analysis of four self-incompatible lines in Brassica napus

Reciprocal hybridization between four self-incompatible lines of Brassica napus: 271, 181, 184 and ‘White Flower’, revealed incompatibility. The reciprocal F₁s obtained by bud pollination showed self-incompatible reactions, and no segregation for self-incompatibility was observed in all the reciprocal F₂ populations, indicating that lines 271, 181, 184 and ‘White Flower’ were genetically identical with regard to self-incompatibility. Observations of self-incompatibility in 17 hybrids from crosses between line 271 and 17 varieties of B. napus showed 10 of the F₁ hybrids to be self-compatible, while four were partially self-compatible and three were self-incompatible. Genetic analysis based on F₂ and BC₁ populations from five self-compatible F₁ hybrids and two self-incompatible F₁ hybrids suggested the existence of at least two loci controlling the self-incompatibility of line 271: one is the S locus, with dominant and recessive relationships between the S alleles, and the other is the suppressor (sp) of the S locus. The sp locus is genetically different from the S locus, and also shows dominant and recessive relationships between the sp alleles.     

Genetic variation and cultivar identification of Jamaican yam germplasm by random amplified polymorphic DNA analysis

Summary We have used random amplified polymorphic DNA (RAPD) analysis to characterize eleven cultivars of the five economically most important yam species grown in Jamaica (Dioscorea alata, D. cayenensis, D. rotundata, D. trifida and D. esculenta). Amplification of genomic DNA samples with nine different arbitrary 10mer primers revealed a total of 338 different band positions, ranging in size from 0.3 to 2.5 kb. RAPD patterns proved to be highly reproducible and somatically stable. While no variation was observed among plants belonging to the same cultivar, a large number of intervarietal and interspecific polymorphisms enabled us to reliably discriminate between all Jamaican cultivars investigated.     

Identification of the series-specific random amplified polymorphic DNA markers of Viola species

Although many Viola species are grown as ornamentals, no information on molecular markers is available for the classification and breeding of Viola. Random amplified polymorphic DNA (RAPD) analysis was applied to identify series-specific molecular markers from the series Pinnatae, Chinensis, and Variegatae of Viola species in the subsection Patellares. We report eight RAPD markers which can be used to distinguish three different series of Viola species based on morphological characters. The eight RAPD markers we identified could be useful to classify Viola species and to assist the hybrid breeding of new varieties.     

Molecular characterization of novel resynthesized rapeseed (Brassica napus) lines and analysis of their genetic diversity in comparison with spring rapeseed cultivars*

Resynthesized (RS) rapeseed generated from interspecific hybridization between suitable forms of Brassica rapa L. (syn. campestris; genome AA, 2n = 20) and B. oleracea L. (CC, 2n = 18) represents a potentially important resource to expand genetic diversity in the narrow gene pool of oilseed rape (B. napus L., AACC, 2n = 38). In this study 165 RS rapeseed lines originating from crosses between an Indian Yellow Sarson (B. rapa ssp. trilocularis) and five different cauliflower (B. oleracea convar. botrytis) cultivars were studied using amplified fragment length polymorphism (AFLP) markers and their genetic diversity was compared in relationship to an assortment of 40 diverse spring oilseed and fodder rape varieties. Using three AFLP primer combinations, a total of 467 polymorphic bands were scored. Cluster analysis allowed differentiation among the different RS lines, which, as expected, were genetically highly divergent from the cultivars. The genetic diversity of the material is discussed in relation to its morphological variability with a view to the implementation of RS lines in oilseed rape breeding.     

Comparisons of Isozyme Patterns in Resynthesized Amphihaploid Rapeseed (Brassica napus) and Their Parental Species Brassica campestris and Brassica oleracea

Resynthesized rapeseed plants produced by embryo culture and the progeny of their selfed parents from Brassica oleracea and Brassica campestris were analyzed by starch gel electrophoresis staining of enzymes GPI, LAP and SDH. In at least one unequivocal case it could be proved for each enzyme system studied that the alleles of both parents arc independently expressed in the newly synthesized individuals and that in the pattern of the dimeric enzyme GPI interlocus bands of the homoeologous loci arc visible. Comparisons with enzyme patterns in cultivated B. napus leads to the conclusion that in general active enzyme loci of both parental genomes are present. This is of great significance in estimating slide frequencies, degree of heterozygosity and other genetic parameters of B. nupus populations/cultivars.     

Genetic diversity of oilseed Brassica napus inbred lines based on sequence-related amplified polymorphism and its relation to hybrid performance

Significant heterosis for seed yield in oilseed rape has created interest in the development of hybrid cultivars. The DNA‐based marker protocol, sequence‐related amplified polymorphism (SRAP) was used to determine genetic diversity among oilseed rape maintainer and restorer lines. This measure was used in an attempt to establish an association between genetic distance and heterosis in hybrids for various agronomic traits. A total of 118 polymorphic loci were generated by 18 SRAP primer combinations. Based on the polymorphism generated by the markers, calculated similarity index values ranged from 0.46 to 0.97. Cluster analysis grouped 10 maintainer and 12 restorer lines into three groups, with the exception of two maintainer lines, PM5 and PM9, which fell outside these groups. The grouping of the lines was largely in agreement with the available pedigree data on their origin and agronomic performance. Analysis of variance among inbred lines and their resulting F₁ hybrids over two locations revealed significant differences for plant height, days to maturity and seed yield, but not for oil content. Substantial mid‐parent heterosis was observed only for seed yield, and ranged from 26% to 169%. All hybrids surpassed their respective inbred lines for this trait, except for a single cross combination of related lines. In general, crosses of lines located in different clusters yielded more than those from the same clusters. Regression analysis revealed a statistically significant relationship between the genetic distance of the parents and seed yield in their hybrid, and their derived mid‐parent and high‐parent heterosis. The correlation coefficient between genetic distance and yield (0.64) indicated a moderately strong relationship, so it is possible that some of the SRAP markers might be linked to quantitative trait loci for seed yield.     

甘蓝型油菜抗寒相关基因的QTL初步定位

低温是影响油菜生产和产量最重要的非生物逆境胁迫因子。为了阐明油菜抗寒性的遗传基础并定位相关的数量性状位点,利用甘蓝型油菜强抗寒GZ恢和弱抗寒10B杂交获得的147个F_2单株为定位群体、以双亲及F2∶3家系为辅助研究材料,利用SSR分子标记结合生理学指标及自然条件下抗寒性调查对抗寒基因QTL进行初步定位。结果表明:630对SSR引物中有160对在双亲间具有多态性,多态率25.40%;160对多态性引物中有102对在F_2群体中有多态性,多态率63.75%,102对多态性引物共检测出多态性位点229个。102个SSR标记构建连锁图,图谱总长974.70 c M,平均距离为5.70 c M。在自然越冬条件下,对F2及F2∶3家系的抗寒性及叶片相对电导率、组织相对含水量、SPAD值、丙二醛(MDA)含量进行测定,分析生理指标与抗寒性的相关性,结果呈显著或极显著相关,表明这些指标可以作为抗寒性评价的参考指标。共定位出5个与抗寒性相关的QTL位点,SPAD值与MDA含量2个指标之间检测到的QTL具有重叠区,这可能是由一因多效或基因紧密连锁引起,可以作为后续分析候选基因的1个热区。为进一步进行甘蓝型油菜抗寒性状相关基因的精细定位及克隆奠定了基础理论。     

Synthesis of low linolenic acid rapeseed (Brassica napus L.) through protoplast fusion

Brassica napus somatic hybrids with low linolenic acid (18:3) content in their seed oil have been produced using fusion partners screened for low 18:3. One somatic hybrid contained only 3.5% 18:3, a level significantly below the mid-parental mean. The low level of 18:3 proved stable in the R₁ generation. Oil content of the lowest 18:3 selection increased from the mid-parental mean (29.3%) in the R₀ generation to 36% in a R₁ field bulk. The R₁ field population also showed some resistance to shattering. This revised version was published online in July 2006 with corrections to the Cover Date.     

甘蓝型油菜抗旱性鉴定研究进展

甘蓝型油菜抗旱育种的最终目的是培育干旱条件下节水、高产的新品种。笔者通过形态及生长发育指标、生理生化指标(光合作用、渗透调节、抗氧化酶活性、内源激素、水分利用率等)、产量和品质指标、综合评价指标等对油菜抗旱性鉴定研究进行了回顾式分析,认为水分胁迫对油菜不同时期形态及生长发育具有重要影响。通过回顾式研究,对甘蓝型油菜抗旱性鉴定研究的现存问题进行分析,并对相关指标与技术展开讨论,以期为油菜抗旱性研究和生产提供参考。同时甘蓝型油菜抗旱性分子机制的阐明必然将油菜抗旱育种研究带入一个崭新的阶段。     

Sexual transfer of Arabidopsis DNA to Brassica napus

Out of 400 reciprocal crosses made between Brassica napus and Arabidopsis thaliana, 200 maternal B. napus flowers produced 31 seeds and 10 seeds developed when A. thaliana was used as the female parent. Of the latter, three seeds successfully germinated but did not develop into mature plants. In contrast, plants from all 32 seeds generated from B. napus were established. DNA from these 31 plants was analysed by 18 probes mapped to the A. thaliana genome, two selectable marker genes and one A. thaliana species-specific repetitive probe. It was found that one plant had identical restriction fragments to A. thaliana with two of the 21 probes used. The gene flow within Brassicaceae is discussed in light of this result.     

甘蓝型油菜遗传多样性及其与杂种表现的关系

以3个双低甘蓝型油菜自交不亲和系、 22个不同来源的父本品种及其按NCⅡ法配制的66个杂种为材料, 应用AFLP及RAPD标记技术研究亲本的遗传多样性及遗传距离与杂种产量、含油量的关系. 结果表明, 根据遗传距离可将3个自交不亲和系分为2组, SI-1300和SI-1310为一组, SI-1320单独为一组; 父本品种分为5组, 中国品种为1组, 国外品     

Identification and inheritance of a partially dominant gene for yellow seed colour in Brassica napus

A yellow‐seeded doubled haploid (DH) line no. 2127‐17, derived from a resynthesized Brassica napus L., was crossed with two black‐seeded Brassica cultivars ‘Quantum’ and ‘Sprint’ of spring type. The inheritance of seed colour was investigated in the F₂, and BC1 populations of the two crosses and also in the DH population derived from the F1 of the cross ‘Quantum’× no. 2127‐17. Seed colour analysis was performed with the colorimeter CR‐300 (Minolta, Japan) together with a visual classification system. The immediate F1 seeds of the reciprocals in the two crosses had the same colour as the self‐pollinated seeds of the respective black‐ and yellow‐seeded female parents, indicating the maternal control of seed colour. The F1 plants produced yellow‐brown seeds that were darker in colour than the seeds of no. 2127‐17, indicating the partial dominance of yellow seed over black. In the segregating BC1 progenies of the two crosses, the frequencies of the black‐ and yellow‐seeded plants fit well with a 1 : 1 ratio. In the cross with ‘Quantum’, the frequencies of yellow‐seeded and black‐seeded plants fit with a 13 : 3 ratio in the F₂ progeny, and with a 3 : 1 ratio in the DH progeny. However, a 49 : 15 segregation ratio was observed for the yellow‐seeded and black‐seeded plants in the F₂ progeny of the cross with ‘Sprint’. It was postulated from these results that seed colour was controlled by three pairs of genes. A dominant yellow‐seeded gene (Y) was identified in no. 2127‐17 that had epistatic effects on the two independent dominant black‐seeded genes (B and C), thereby inhibiting the biosynthesis of seed coat pigments.     

Linkage of random amplified polymorphic DNA markers to downy mildew resistance in cucumber (Cucumis sativus L.)

Marker assisted selection (MAS) may improve the efficiency of breeding downy mildew resistant cucumber cultivars. A study was conducted to identify random amplified polymorphic DNA (RAPD) markers linked to the downy mildew resistance gene (dm) which would be suitable for MAS. A total of 145 F₃ families from two populations (55 from the WI 1983G × Straight 8 population and 90 from the Zudm1 × Straight 8 population) were evaluated over five locations in North America and Europe. Resistant and susceptible F₃ families were identified and mean family resistance ratings were used to type individual F₂ plants. No evidence for race differences in the pathogen (Psuedoperonospora cubensis (Berk. & Curt.) Rostow) between North America and Europe was found. Phenotypic correlations between locations ranged from 0.3 to 0.7. Of the 135 polymorphic RAPD markers identified from 960 primers, five were linked to dm - G14₈₀₀, X15₁₁₀₀, AS5₈₀₀, BC519₁₁₀₀, and BC526₁₀₀₀. In the WI 1983G × Straight 8 population, G14₈₀₀ was linked to dm at 16.5 cm, AS5₈₀₀ at 32.8 cm, BC519₁₁₀₀ at 9.9 cm, and BC526₁₀₀₀ at 19.2 cm. In the Zudm1 ×Straight 8 population, G14₈₀₀ was linked at 20.9 cm, X15₁₁₀₀at 14.8 cm, AS5₈₀₀ at 24.8 cm, and BC526_{_1000} at 32.9 cm. MarkersG14₈₀₀ and BC519₁₁₀₀ were linked in repulsion to the dm allele, and X15₁₁₀₀, AS5₈₀₀, and BC526₁₀₀₀ were linked in coupling phase. These genetic markers may be exploited to develop an efficient MAS strategy for breeding resistant cucumber cultivars. This revised version was published online in July 2006 with corrections to the Cover Date.