Further Work on Plant Injection for Diagnostic and Curative Purposes

Summary

Randomly Amplified Polymorphic DNA (RAPD) markers were used to evaluate genetic similarity and inter-relationship among31 acid citrus species and cultivars, including sour oranges (six accessions); ‘Yuzu’ (four accessions) andits relatives (21 accessions). Out of the 60 decamer primers screened, 27 were selected which produced 108 markers; 76 of which were polymorphic. Species or cultivar-specific RAPD markers were also found. A dendrogram based on genetic distance implied that sour oranges were very distinct from ‘Yuzu’ and its relatives. ‘Yuzu’ accessions were very closely linked to each other, however; for the other specimens genetic polymorphism could easily be detected by RAPDs and the genetic variation between accessions was quite high and revealed their different origins. In this study some RAPDs allowed the distinction of very close cultivars, for instance ‘Kabosu’ from ‘Aka kabosu’.     

菠萝蜜遗传多样性的ISSR分析

用ISSR标记方法对76份菠萝蜜(Artocarpus heterophyllus Lam.)种质资源DNA的遗传多样性进行了检测,24个引物共检测到477条带,其中427条具多态性(占89.52%),平均PIC为0.23,24个引物能把76份种质完全区分开来。遗传距离分析结果显示供试种质的遗传多样性较低,在DNA水平上的遗传相似系数为0.626～0.945,平均为0.775。聚类分析表明,76份菠萝蜜材料在遗传距离系数为0.752处可分为4大类,其中热带植物园的种质WJ2和GSYWJ1与其他种质的遗传距离相对较大,干胞和湿类型不能独立聚类。另外各地区的种质与雷州半岛种质明显分开聚类,而雷州半岛各地区的种质混杂聚类在一起,不能按地区单独聚类。     

SSR-based DNA fingerprints reveal the genetic diversity of Sicilian olive (Olea europaea L.) germplasm

Summary

Twelve published simple sequence repeat (SSR; microsatellite) markers, belonging to the ssrOeUA-DCA, GAPU and UDO series, were tested in a panel of 46 accessions of olive germplasm belonging to 30 unique cultivars collected in seven Provinces of Sicily. Four well-known reference olive cultivars were also added. The analysis was carried out on an automatic capillary sequencer using fluorescent dyes, and fragment sizes were determined using internal standards. The results allowed us to rank the SSRs assayed according to their information content and reproducibility. Up to 115 alleles were identified (119, if those unique to sport mutations were included), the frequency of which allowed genetic relationships among accessions to be investigated. The probability that two unrelated genotypes displayed the same SSR pattern at all loci examined was calculated to be as low as 1.18 10^–11. Sixteen accessions were identified as synonyms. Of these, eight matched perfectly with another accession at all SSR loci examined. The others showed one or two allelic differences from the reference accession. These were interpreted as mutations. Otherwise, all accessions were clearly separated from each other. Two likely parentages were also identified (‘Giarfara’ = ‘Nocellara del Belice’

‘Cacaridduni’; and ‘Pizzo di Corvo’ = ‘Nocellara Etnea’ ‘Tonda Iblea’). The genetic diversity of the pool represented by the unique accessions was very high, reflecting the richness of the olive germplasm accumulated in Sicily. A database of the accessions is available to the scientific community (http://www.unipa.it/germolive/ssr.html) to facilitate comparisons of data.     

雷州半岛菠萝蜜种质遗传多样性的RAPD分析

 用RAPD标记方法对我国雷州半岛的65份菠萝蜜(Artocarpus heterophyllus Lam. ) 实生种质资源DNA的遗传多样性进行了检测, 16个引物共检测到78条带, 其中69条具多态性(占88.4% ) 。聚类分析表明, 65份菠萝蜜材料在遗传距离0.26处可分为3大类。供试种质虽具有丰富的形态性状变异, 但在DNA水平上平均相似性系数为0.7341。干、湿胞类型及引自马来西亚的种质均不能独立聚类。对菠萝蜜品种的干、湿胞分类法及菠萝蜜品种的引种区域进行了讨论。     

Analysis of genetic diversity among Indian short-day onion (Allium cepa L.) cultivars using RAPD markers.

Summary

Randomly amplified polymorphic DNA (RAPD) markers were used to estimate genetic diversity among 24 cultivars of short-day onions. Total genomic DNA was extracted and subjected to RAPD analysis using 90 arbitrary 10-mer primers. Of these, 15 primers were selected which yielded 137 bands, 91.24% of which were polymorphic. None of the primers produced a unique banding pattern for each cultivar. RAPD data were used to calculate a Squared-Euclidian Distance matrix which revealed a minimum genetic distance between cultivars ‘AFLR-722’ and ‘PBR-140’, and a maximum genetic distance between cultivars ‘PBR-139’ and ‘A.Kalyan’, and ‘MS-48’ and ‘A.Kalyan’. Based on the distance matrix, cluster analysis was done using a minimum variance algorithm.The dendrogram thus generated, based on Ward’s method, grouped the 24 onion cultivars into two major clusters. The first cluster consisted of cultivars from the northern region, and the second of cultivars from the southern region of India. The present study shows that there is high diversity among the onion cultivars selected and indicates the potential of RAPD markers for identification and maintenance of onion germplasm for crop improvement purposes.     

基于叶绿体DNA信息的南方梨属种质的遗传多样性和演化分析

选取5个叶绿体DNA非编码区trnL-trnF-1、trnL-trnF-2、trnS-psbC、accD-psaI、rps16-trnQ和1个基因区域rbcL,对中国秦岭淮河以南地区的梨（Pyrus L.）资源进行遗传多样性和演化分析。将188份梨资源的测序结果整合得到长度为5 612 bp（包含缺失片段）,共检测出13个单倍型、4个单一突变位点、16个简约性信息位点和14个插入/缺失片段（InDels）,单倍型多样性Hd为0.7178,核苷酸多样性为0.35 × 10-3。accD-psaI区域的分析结果显示该区域的Hd值最高,为0.7177;rbcL区域的Hd值最低,为0.0317。其中162份梨资源检测出12个单倍型,Hd为0.713,核苷酸多样性为0.29 × 10-3;梨的不同地理群体中湖南省的10个梨地方品种的Hd最高,为0.889;来自贵州省的8个品种的Hd最低,为0。138份砂梨地方品种共检测出8个叶绿体单倍型,其中单倍型H2、H4和H5在130份砂梨和37份白梨品种中检测到,单倍型H6、H10、H11、H12和H13在8份砂梨地方品种‘青皮钟’、‘甜宵梨’、‘细花红梨’、‘惠阳红梨’、‘麝香梨’、‘塘湖青’、‘红粉’和‘横县浸泡梨’中检测到。Median-joining Network图显示单倍型H6和H11为古老单倍型,H1、H4和H5为较进化的单倍型。根据叶绿体DNA序列变异信息可推测秦岭淮河以南地区的砂梨和白梨亲缘关系较近,同时来自湖南地区的砂梨种质资源具有丰富的遗传多样性。     

Isolation and characteristics of eight novel polymorphic microsatellite loci from the genome of garlic (Allium sativum L.)

Here we characterized eight novel polymorphic SSR markers, developed from an enriched genomic library of garlic (Allium sativum L.). These SSRs produced a total of 64 alleles across 90 garlic accessions, with an average of 8 alleles per locus. Values for observed (H_O) and expected (H_E) heterozygosity ranged from 0.16 to 0.77 (mean = 0.44) and from 0.22 to 0.86 (mean = 0.65), respectively. Six loci deviated significantly (P < 0.05) from Hardy–Weinberg equilibrium (HWE). The averages of gene diversity and PIC values were 0.65 and 0.62, respectively. The mean genetic similarity coefficient was 0.4380, indicating that among garlic accessions existed wide genetic variation. Based on 64 alleles obtained by 8 SSRs, a phenogram was constructed to understand the relationships among the 90 accessions. These newly developed SSRs should prove very useful tools for genotypes identification, assessment of genetic diversity and population structure in garlic.     

Molecular diversity of Chinese Cucurbita moschata germplasm collections detected by AFLP markers

Cucurbita moschata is an important vegetable crop. Although a total of 1032 landraces of C. moschata are maintained in China, little is known about their genetic diversity. Molecular characterization is needed to facilitate the use of this C. moschata germplasm collection in breeding. Seventy-four Chinese accessions and 15 accessions from other countries were selected for evaluation based upon variation in fruit traits and geographical origin of molecular diversity with AFLP analysis. Nine pairs of EcoRI/MseI primers produced 500 fragments, of which 75.57% were polymorphic, indicating a high degree of diversity. The accessions from China were classified into two clusters, which were clearly differentiated from the accessions originating from Mexico, Guatemala, Honduras, and Ecuador. Chinese group genetically more closely related to other Asian countries group (India and Japan). In general, the accessions from the Americas had a greater number of unique loci than those from China. The differences are probably due to a limited number of introductions and genetic drift. The Americas are the center of origin of C. moschata and therefore more diverse. With AFLP analysis, the accessions did not clearly group according to fruit shape; however, sub-clusters exist in acorn- and dumbbell-shaped accessions. The assessment of genetic distance, along with some unique traits among the different genotypes, could be useful in further genetic studies and the selection of the most adequate accessions for use in breeding programs.     

Jasmonic acid effects on gibberellic acid-induced seedlessness in ‘Neo Muscat’ table grapes

Summary

The effect of jasmonic acid (JA), applied alone or in combination with gibberellin A3 (GA₃), on the induction of seedlessness in ‘Neo Muscat’ (Vitis vinifera) was investigated. Endogenous JA levels in the florets of ‘Neo Muscat’ and ‘Delaware’ (Vitis vinifera x Vitis labrusca) were also studied. The proportion of seedless berries induced by application of GA₃ alone 12 d before full bloom (DBB) was 17.0%, and at 6 DBB it was 62.7%. The proportion of seedless berries induced by GA₃ + JA applied 6 DBB increased significantly with increasing JA concentration, while pollen germination decreased quadratically with increasing JA concentration. JA did not affect seed number per berry or the seed’s fresh weight. Treatment with JA alone also induced seedlessness effectively. Although the fresh weight of the seedless berries induced by the application of GA₃ + JA 12 DBB decreased quadratically with increasing JA concentration, there were no significant differences between the treatments at 6 DBB. Fruit soluble solids and titratable acidity were not affected by JA. There were no obvious differences in the amounts of JA in the florets of ‘Delaware’ and ‘Neo Muscat’.     

Development of new microsatellite markers for molecular diversity analysis of Citrus species

Summary

Thirty microsatellite loci for further genetic analysis of Citrus species were developed by constructing a microsatellite-enriched library using capture with streptavidin-coated magnetic beads and used to assess genetic diversity in 40 Citrus accessions. In total, 150 alleles were detected, with an average of five alleles per locus. The average gene diversity and polymorphism information content values were 0.58 and 0.52, and ranged from 0.35 – 0.74 and from 0.32 – 0.70, respectively. Values for the observed and expected heterozygosities ranged from 0.13 – 1.00 and from 0.36 – 0.75, respectively. Fifteen loci deviated from the Hardy-Weinberg equilibrium (P < 0.05). The mean similarity coefficient among accessions was 0.5906. Based on the UPGMA algorithm, two main groups were successfully identified. These new microsatellite markers can be used to further investigate the genetics of, and phylogenic relationships in Citrus spp.     

Effect of pruning and plant spacing on the growth of cherry rootstocks and their influence on stem water potential of sweet cherry trees

Summary

The aims of this work are to describe the effects of pruning and planting density on growth and water relations of ungrafted and grafted sweet cherry trees. A trial with cherry rootstocks ‘Prunus avium’, ‘CAB 11E’, ‘Maxma 14’, ‘Gisela 5’ and ‘Edabriz’ was begun in 1997. Pruning severities were applied to the rootstocks (0, 30, 60 and 90% of the vegetative growth was removed corresponding to P1, P2, P3 and P4 treatments, respectively) after planting to two plant spacings (S1 = 0.25 × 1.0 m and S2= 0.45 × 1.5 m). Canopy, root growth and leaf water potential (ψ_leaf) were quantified throughout the growing season. Pruning significantly affected root length and root weight of the rootstocks. Uncut plants (P1) showed a heavier and expanded root biomass (231 g and 108 m) than the intensively pruned plants (P4) (187 g and 75 m). The greater root biomass was obtained with the spacing/pruning combination, S1/P1 (285 g), and the smaller with S1/P4 (180 g) and S2/P4 (176 g). ψ_leaf varied significantly between the rootstocks and plant spacing but not with pruning. ‘Maxma 14’ and ‘P. avium’ attained the lowest values of midday ψ_leaf, –2.28 and –2.04 MPa, but the highest values of predawn ψ_leaf, –0.29 and –0.25 MPa, respectively. Generally, with high density (S1), the rootstocks exhibited lower predawn and midday ψ_leaf. In 1998, cultivars ‘Burlat’, ‘Summit’ and ‘Van’ were grafted onto rootstocks and a trial was installed in 1999. Predawn and midday stem water potential (ψ_stem) on cherry trees, measured in 2002, were affected significantly by the rootstock/genotype combination. Cultivars grafted on ‘P. avium’ and ‘Maxma 14’ showed the less negative midday ψ_stem, –1.36 and –1.42 MPa respectively, so these rootstock genotypes perhaps induced a higher drought resistance to the scion. Recorded data show that the scion-rootstock interaction with regard to production performance under water deficits may be an important consideration in cherry tree planting strategies.     

Genetic diversity in fruiting-mei,apricot, plum and peach revealed by AFLP analysis

Summary

High-throughput amplified fragment length polymorphism (AFLP) analysis, employing a two-dye automated DNA sequencer, was used to genotype accessions of four important Prunus spp. fruit crops in China, fruiting-mei (P. mume), apricot (P. armeniaca), plum (P. salicina, and P. ussuriensis) and peach (P. persica). Nine primer pairs generated a total of 2,089 AFLP bands, of which 94.4% were polymorphic. Levels of polymorphism and the genetic relationships among the crops were studied.The fixation index (F_st) of the four different fruit trees ranged from 0.53 – 0.82, and the average genetic diversity (AGD) in these species ranged from 3.2% to 12.8%. These four stone fruit species could be clustered into four clear groups on a phylogenetic tree based on bootstrap analysis. Phylogenetically, fruiting-mei was shown to be closest to apricot and most distant from peach.     

Genetic diversity of Dimocarpus longan in China revealed by AFLP markers and partial rbcL gene sequences

Amplified fragment length polymorphism (AFLP) and partial rbcL gene sequencing were used to investigate genetic diversity among various longan (Dimocarpus longan Lour) accessions as well as a presumed closely related species Dimocarpus confinis How et Ho and litchi (Litchi chinensis Sonn). No significantly shared AFLP fragment was found between the three species, indicating that D. confinis and litchi are very far in genetic distance from any longan accession studied. Partial rbcL sequences of 501 bp from the first coding site in these species were obtained, which revealed several substitutes. One such DNA base pair substitute resulted in an amino acid difference between longan and litchi. Furthermore, another 4 bp resulted in a two amino acid difference between longan and D. confinis, which was consistent with AFLP results and indicated that D. confinis should be excluded from the longan genus, Dimocarpus. Within the longan species, no DNA substitute was found. Using nine primer combinations, a total of 66 AFLP markers were obtained from 41 longan accessions. One non-Chinese longan accession ‘Miaoqiao’ was distinctly different from all other longan cultivars collected in China, indicating that more genetic resources of longan might be collected also from longan production regions outside of China. AFLP markers might be developed to identify longan cultivars as well as expedite progeny screening in breeding programs of this perennial fruit tree.     

Assessment of genetic variability in Italian heritage peach resources from Emilia-Romagna using microsatellite markers

Summary

Heritage peach (Prunus persica L. Batsch) varieties in Emilia-Romagna, Italy’s leading peach-producing region, are largely marked by melting, juicy, aromatic white-fleshed fruit with a short shelf-life and susceptibility to bruising.While these varieties have rapidly been replaced since the 1950’s by yellow-fleshed peaches from US breeding programmes with improved handling resistance, the Fruit Research Unit of the Agricultural Research Council, at Forlì, has begun an effort to safeguard and characterise the heritage varieties of Emilia-Romagna. The programme has screened 26 local heritage accessions using a set of 16 highly polymorphic microsatellite (simple sequence repeat; SSR) markers to assess genetic diversity, to elucidate inter-relationships, and to resolve any cases of homonymy. Seven international peach cultivars have been included in order to standardise allele scoring. This study resulted in 19 unique molecular profiles among the heritage accessions, and a relatively high mean observed heterozygosity value of 0.49 for a germplasm pool from a restricted region, an indicator of potential genetic diversity. The main morphological features of the heritage peaches are reported, and potential benefits for future breeding programmes discussed.     

Maintaining product quality in raisins throughout the introgression of powdery mildew resistance

The availability of powdery mildew-resistant raisin accessions will lower grower production costs and enhance the environment through reduced fungicide use. To achieve this objective, backcrossing has been employed with Vitis romanetii as the source of non-race-specific powdery mildew resistance. Initial crosses of powdery mildew-resistant F₁ hybrid B36-45 with seeded raisin cultivars ‘Rangspray’ and ‘Raisin de Palestine’ yielded seedless powdery mildew-resistant first-generation backcross selections C87-41 and C87-106, which were used to develop second-generation backcross populations. Principal component analyses consistently identified ‘wrinkle,’ ‘meatiness,’ ‘product attractiveness’ and ‘skin persistence’ as being the quality characteristics most important in discriminating among powdery mildew-resistant raisin accessions and commercial raisin cultivars. Raisin quality ratings were much improved across most evaluated characters in second-generation backcross families as compared with parental powdery mildew- resistant accessions C87-41 and C87-106. After two generations of backcrossing, powdery mildew-resistant raisin selections were identified with product quality characteristics similar to those of commercial raisin cultivars.     

Genetic relatedness (diversity) and cultivar identification by randomly amplified polymorphic DNA (RAPD) markers in teasle gourd (Momordica dioica Roxb.)

Genetic relationship and variation of 29 accessions of teasle gourd (Momordica dioica Roxb.) and 1 accession of Momordica cochinchinensis Spreng. (wild relatives of teasle gourd) were examined by RAPD analysis using 44 dodecamer oligonucleotide primers. A total of 496 fragments were produced by 44 primers of which 95% bands were polymorphic. Using presence or absence of specific RAPD markers or combination of primers, 23 out of 30 accessions were identified. The genetic relatedness or genetic distance based on Nei and Li's genetic similarity varied from 0.86 to 0.65 with an average of 0.74 among 29 M. dioica accessions (when M. cochinchinensis excluded). In the phenetic dendrogram developed from cluster analysis using UPGMA method, M. cochinchinensis was out grouped as single accession, while others showing relatively weak grouping formed four groups. Clustering pattern did not demonstrate any relationship between geographical origin and genetic diversity. A DNA extraction method has been standardized. This is the first report of using RAPD techniques in teasle gourd. It was concluded that RAPD analysis is a useful tool for genotypic identification and estimation of genetic similarity in teasle gourd.     

Wide genetic diversity of Rosa damascena Mill. germplasm in Iran as revealed by RAPD analysis

The genetic relationships among 41 Rosa damascena accessions from various cultivation areas of Iran and one accession from Bulgaria were analyzed using 31 RAPD (Randomly Amplified Polymorphic DNA) primers. Each primer exhibited 3–12 banding patterns for a total of 343 scorable and 184 polymorphic bands. The combination of 11 primers was found optimal for discrimination of 42 accessions with very low values of cumulative confusion probability (9.7 × 10⁻⁵); indicating that only one pair from over 10,000 distinct pairs of accessions would be indistinguishable. Unweighted pair group method cluster analysis based on similarity values revealed 10 groups at the distance of 0.85. The Bulgarian genotype grouped with the majority of the Iranian genotypes in a main cluster. Results of molecular variance analysis (AMOVA) indicated that the major proportion (65.7%) of the total genetic variation was within collecting provinces rather than between them. The wide genetic variation seen for R. damascena in Iran indicates that Iran is a center of genetic diversity for this species and that there is a promising future for the breeding.