AFLP analysis of genetic diversity within and between populations of perennial ryegrass (Lolium perenne L.)

Amplified fragment length polymorphism (AFLP) analysis has been used to measure genetic diversity in perennial ryegrass (Lolium perenne L.) and to relate intra- and interpopulation variation to breeding history. Cluster analysis of AFLP data from contrasting populations showed features consistent with the origins of these varieties. Significant differences in intrapopulation diversity were detected and partial separation of different cultivars was observed. Restricted base cultivars, derived from small numbers of foundation clones, were suitable for this type of study, allowing near complete discrimination of closely related cultivars. Analysis of bulked samples was based on the pooling of genomic DNA from 20 individuals from 6 selected populations. Cluster analysis of AFLP data from bulked samples produced a phenogram showing relationships consistent with the results of individual analysis. AFLP profiling provides an important tool for the detection and quantification of genetic variation in perennial ryegrass. This revised version was published online in August 2006 with corrections to the Cover Date.     

用RAPD、ISSR和AFLP标记分析系谱关系明确的甘薯品种的亲缘关系

用RAPD、ISSR和AFLP标记对系谱关系明确的7个甘薯品种进行了亲缘关系分析。24个RAPD引物、14个ISSR引物和9对AFLP引物分别扩增出173、174和168条多态性带。3种分子标记在检测甘薯品种间遗传差异上相关程度高，其中RAPD与ISSR之间的相关系数最大为0.9328。用ISSR标记估计的品种间遗传距离为0.1286~1.0932，平均0.4883，大于其余2个标记的估计值。3种分子标记皆可揭示甘薯品种的亲缘关系，其中ISSR标记产生的聚类图与系谱图最吻合，认为ISSR标记更适于分析甘薯品种的亲缘关系。     

Study on Somaclonal Variation through a Callus Regeneration System in Betula Platyphylla

Calli have been widely used as research materials for clonal propagation and transformation of trees owing to their high multiplication rates and capacity of large-scale propagation. Till now, very little is known on the somaclonal variation in Betula platyphylla culture, and especially, in regenerated plants （Schween et al., 2003; Nakano et al., 2003）.     

Molecular mapping of a dominant genic male sterility gene Ms in rapeseed (Brassica napus)

Rs1046AB is a genic male sterile two‐type line in rapeseed that has great potential for hybrid seed production. The sterility of this line is conditioned by the interaction of two genes, i.e. the dominant genic male sterility gene (Ms) and the suppressor gene (Rf). The present study was undertaken to identify DNA markers for the Ms locus in a BC₁ population developed from a cross between a male‐sterile plant in Rs1046AB and the fertile canola‐type cultivar ‘Samourai’. Bulked segregant analysis was performed using the amplified fragment length polymorphism (AFLP) methodology. From the survey of 480 AFLP primer combinations, five AFLP markers (P10M13₃₅₀, P13M8₄₀₀, P6M6₄₁₀, E7M1₂₃₀ and E3M15₁₀₀) tightly linked to the target gene were identified. Two of them, E3M15₁₀₀ and P6M6₄₁₀, located the closest, at either side of Ms at a distance of 3.7 and 5.9 cM, respectively. The Ms locus was subsequently mapped on linkage group LG10 in the map developed in this laboratory, adding two additional markers weakly linked to it. This suite of markers will be valuable in designing a marker‐assisted genic male sterility three‐line breeding programme.     

Incidence and molecular markers of 2n pollen in Populus tomentosa Carr.

Microscopic examination, amplified fragment length polymorphism (AFLP) and SCAR (sequence-characterized amplified region) molecular markers were employed to determine the incidence of 2n pollen (unreduced pollen) in Chinese white poplar (Populus tomentosa Carr.) and to identify related molecular markers. The presence of a parallel and tripolar spindle at metaphase II and the absence of cytokinesis at telophase II were found to be determining factors in 2n pollen formation. A group of 298 clones that originated from their indigenous areas were investigated for the production of 2n pollen based on pollen size differences, both within a clone and between n and 2n pollen. Pollen grains were collected from 224 of the clones, six of which were subsequently determined to produce only normal pollen; the remainder produced 2n pollen at different frequencies (0.6–21.9%). The frequency at which 2n pollen was produced was significantly and highly significantly different among and within indigenous populations, respectively. Clones produced by the six normal and twenty-two 2n pollen clones were selected for AFLP analysis. Following an initial screening with 55 primer combinations, the E50-M38 (CAT/ACT) primer was identified: it generated a PCR fragment (246 bp) from the normal clones, but not from the 2n pollen producers. In addition, the E31-M50 (AAA/CAT)-amplified DNA fragment (204 bp) was present in 2n pollen producers, and absent in normal clones. These two discriminating AFLP markers were developed into easily detectable SCAR (sequence characterized amplified region) markers which can be used in combination with previously developed AFLP markers to distinguish between normal and 2n pollen clones.     

Variation of the genomic proportion of the recurrent parent in BC1 and its relation to yield performance in sorghum (Sorghum bicolor) breeding for low-input conditions

In order to define the variation of the genomic proportion of the recurrent parent [G(RP)] and its relation to yield, G(RP) of individual BC₁ plants of two sorghum populations composed of a high‐yielding cultivar as recurrent parent (RP) and a donor with superior drought resistance or grain quality, respectively, was estimated using AFLPs and SSRs. G(RP) in BC₁ ranged from 0.53 to 0.95 and averaged to 0.76 in the population (NP4453 × ‘SV‐2’) × ‘SV‐2’. G(RP) varied between 0.60 and 0.86 and averaged to 0.74 in the BC₁ of (ICV‐219 × ‘SV‐2’) × ‘SV‐2’. Results show that plants with a G(RP) equivalent to BC₂ (0.875) or BC₃ (0.938), respectively, can be selected from BC₁. Yield performance of BC₁S₁ families was tested in field trials carried out in South Africa. The correlation between yield and G(RP) in BC₁ was low. Selection according to G(RP) did not result in an effective preselection for yield.     

Interspecific hybrids of sunflower as a source of Sclerotinia resistance

Sclerotinia sclerotiorum is considered the most devastating pathogen of sunflower grown in humid environments. In this study, progenies of partial hybrids between Helianthus maximiliani, a wild species that has been shown to be resistant to S. sclerotiorum, and H. annuus were characterized by amplified fragment length polymorphism (AFLP) analysis to identify whether there are introgressions from H. maximiliani into the cultivated sunflower at the molecular level. Wild species‐specific fragments as well as fragments not found in either parent were detected. Progenies tended to cluster together according to the original partial hybrids in the dendrogram by the use of bootstrap procedures. The progenies were studied for their reaction to S. sclerotiorum using artificial inoculation of sunflower heads. Some of the progenies showed a higher level of resistance compared with resistant inbred lines. It was possible to identify two AFLP‐fragments which seem to be linked to Sclerotinia resistance.     

Vine-1 retrotransposon-based sequence-specific amplified polymorphism for Vitis vinifera L. genotyping

The sequence‐specific amplification polymorphism (S‐SAP) method, derived from the amplified fragment length polymorphism (AFLP) technique, produces amplified fragments containing retrotransposon long terminal repeat ( LTR ) sequence at one end and a host restriction site at the other. The development and application of this procedure to the LTR of the Vine‐1 element from grapevine is reported. Two primers derived from one of the LTR sequences flanking the retrotransposon were used in combination with MseI degenerated primers on 15 grapevine accessions. S‐SAP results were compared with AFLP data. The heterozygosity and gene diversity values were higher for S‐SAP than for the AFLP procedure. Results show that S‐SAP amplification is effective in identifying polymorphisms and defining genetic distances among cultivars, and could be used for fingerprinting and for ‘Traminer’ clone identification. To the contrary Vine‐1 retrotransposon‐based S‐SAP was not able to distinguish ‘Pinot’ clones.     

Genetic mapping of AFLP markers in Japanese bunching onion (<Emphasis Type="Italic">Allium fistulosum</Emphasis>)

Summary The first genetic linkage map of Japanese bunching onion (Allium fistulosum) based primarily on AFLP markers was constructed using reciprocally backcrossed progenies. They were 120 plants each of (P₁)BC₁ and (P₂)BC₁ populations derived from a cross between single plants of two inbred lines: D1s-15s-22 (P₁) and J1s-14s-20 (P₂). Based on the (P₂)BC₁ population, a linkage map of P₁ was constructed. It comprises 164 markers – 149 amplified fragment length polymorphisms (AFLPs), 2 cleaved amplified polymorphic sequences (CAPSs), and 12 simple sequence repeats (SSRs) from Japanese bunching onion, and 1 SSR from bulb onion (A. cepa) – on 15 linkage groups covering 947 centiMorgans (cM). The linkage map of P₂ was constructed with the (P₁)BC₁ population and composed of 120 loci – 105 AFLPs, 1 CAPS, and 13 SSRs developed from Japanese bunching onion and 1 SSR from bulb onion – on 14 linkage groups covering 775 cM. Both maps were not saturated but were considered to cover the majority of the genome. Nine linkage groups in P₂ map were connected with their counterparts in P₁ map using co-dominant anchor markers, 13 SSRs and 1 CAPS.     

Marker assisted polycross breeding to increase diversity and yield in perennial ryegrass (Lolium perenne L.)

Summary In this study, the application of molecular markers to optimise genetic diversity in a polycross breeding program of perennial ryegrass (Lolium perenne L.) was evaluated. The genetic diversity among 98 potential parental plants from three maturity groups (early, intermediate and late flowering) was investigated using amplified fragment length polymorphism (AFLP) markers. For each maturity group, two polycrosses of six parental plants with contrasting levels of genetic diversity were composed. Average genetic diversity among parents selected for narrow polycrosses was 36% lower than among parents selected for wide polycrosses. Diversity within first generation synthetic progenies (Syn1) was proportional to the diversity within the respective parental polycrosses. However, differences were less pronounced with Syn1 progenies from narrow polycrosses showing 16% reduced diversity when compared to Syn1 progenies from wide polycrosses. Multivariate analyses allowed for a clear separation of the six Syn1 progenies based on AFLP markers and demonstrated their genetic distinctness. Evaluation of dry matter yield, date of ear emergence and stem length of Syn1 and Syn2 progenies showed progenies from wide polycrosses to be constantly higher yielding when compared to progenies from narrow polycrosses. On the other hand, there was no significant difference in variability for the two morphological traits between progenies of narrow- and wide polycrosses. The results presented here provide evidence for an efficient application of molecular markers to select genetically diverse polycross parents which resulted in an average yield increase of 3.8%.