Chloroplast DNA diversity in brinjal eggplant (Solanum melongena L.) and related species

Chloroplast DNA (cpDNA) samples of brinjal eggplant (S. melongena) and representative related species including S. incanum sensu lato (or S. campylacanthum sensu stricto), S. lichtensteinii, S. marginatum, S. macrocarpon, S. anguivi and S. aethiopicum and also S. nigrum as an outgroup taxon, were digested by 14 restriction enzymes and analyzed by using electrophoresis and a cpDNA probe. All the species used here were clearly separated in the cpDNA analysis, except the pair S. anguivi and S. aethiopicum. From the dendrogram constructed by the unweighted pair-group method, it is suggested that S. incanum is the closest to S. melongena and the next closest species is S. macrocarpon followed by S. aethiopicum (and S. anguivi), S. lichtensteinii, S. marginatum and finally the outgroup taxon S. nigrum. The tree derived by the neighbour-joining method suggests phyletic relationships that agree with those indicated by crossability and seed coat anatomy, but conflict with conventional classifications based on morphology. In particular, members of sections Oliganthes and Melongena are not separated and no cpDNA variation was found within either of the morphologically diverse cultigens, S. aethiopicum and S. melongena. Paradoxically, the morphologically similar species S. incanum, S. lichtensteinii and S. marginatum have diverged greatly in their cpDNA. The significance of these results is discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Chloroplast DNA analysis of eggplant (Solanum melongena) and related species for their taxonomic affinity

Summary Total chloroplast DNA (cpDNA) from Solanum incanum, a wild relative of eggplant, was used to probe total DNA of Solanum melongena (eggplant). The DNA fragments detected were the same as observed using purified chloroplast DNA. Chloroplast DNAs were also analysed for nine species of Solanum that are cross-compatible with eggplant: S. aethiopicum, S. anguivi, S. gilo, S. incanum, S. indicum, S. integrifolium, S. macrocarpon, S. olivare and S. panduriforme.Restriction fragments generated by eight enzymes were recorded as present or absent, and a matrix for all fragment positions, species and enzymes was used for cluster analysis. In the resulting dendrogram, the species tested formed three distinct groups: (1) S. aethiopicum, S. anguivi, S. gilo, S. indicum, S. integrifolium and S. olivare, (2) S. incanum, S. melongena and S. panduriforme, (3) S. macrocarpon. Six species of the first group belonging to section Oliganthes appears more closely related to the second group members belonging to section Melongena than does S. macrocarpon, which also belongs to section Melongena. Within the second group, S. panduriforme is slightly more like eggplant than is S. incanum.     

PCR-RFLP analysis of the whole chloroplast DNA from three cultivated species of cotton (Gossypium L.)

Variations of the chloroplast DNA (cpDNA) from three of the four cultivated species of cotton (Malvaceae); Gossypium barbadense L., Gossypium hirsutum L., Gossypium arboreum L. and its synonym Gossypium nanking Meyen., were analyzed. Using specific set of primers, the whole circular cpDNAs from the four test species were amplified. These were subsequently digested with the use of seven restriction enzymes. The amplified fragments of the whole cpDNAs of the diploid cultivated cotton G. arboreum and its synonym G. nanking did not show any differences. However, the allotetraploid cultivated cottons G. barbadense and G. hirsutum, showed some fragment length differences directly visible after amplification and two types of restriction fragment length polymorphism (RFLP), the first appeared as slightly lengthened bands and the other as gain or loss of a restriction site. The results also showed that the chloroplast genomes of the allotetraploid cultivated cottons are highly similar to the diploid cultivated cottons tested in terms of length and digestion patterns. The detected amplified length differences, RFLPs and the restriction sites can be considered as species specific markers for the allotetraploid cultivated cottons, which could be a useful tool for future studies of the cpDNA of the genus Gossypium L.     

Interspecific hybrids between three eggplant (Solanum melongena L.) cultivars and two wild species (Solanum torvum Sw. and Solanum sisymbriifolium Lam.)

Three Greek eggplant cultivars, ‘Langada’, ‘Tsakoniki’ and ‘Emi’ (2n= 24), were crossed with two wild species (Solanum torvum Sw., 2n= 24 and Solanum sisymbriifolium Lam., 2n= 24). Ovules isolated 15-27 days after pollination were cultured in a modified MS medium at 24°C and a 16h photoperiod. Fifty days later, the ovules were dissected and the interspecific embryos were cultured in the same medium. Interspecific hybrids were achieved only from crosses between the eggplant cultivars and S. torvum. The hybridity of the putative interspecific F¹ hybrid (Solanum melongena×S. torvum) was confirmed by using morphological and biochemical (isozyme isocitrate dehydrogenase A, phosphoglucomutase A, phosphoglucose isomerase B, 6-phosphogluconate dehydrogenase A, 6-phosphogluconate dehydrogenase B) markers. The F¹ plants (‘Langada’×S. torvum) were selfpollinated and backcrossed to both parents. Fruits, however, were produced only when the F¹ hybrid was backcrossed as female with the eggplant cultivar ‘Langada’.     

低温胁迫对茄子幼苗叶片光合特性的影响

以茄子702幼苗为试材,研究了低温胁迫对茄子幼苗光合特性的影响.结果表明,低温胁迫下,茄子幼苗的叶绿素含量、叶绿素a/b、净光合速率(Pn)、蒸腾速率(Tr)、气孔导度(Gs)、光饱和点(LSP)、光饱和光合速率、表观量子效率(AQY)、CO2饱和点(CSP)、羧化效率(CE)和RuBP最大再生速率均下降;光补偿点(LCP)、CO2补偿点(CCP)均上升;胞间CO2浓度(Ci)先降低后升高.以上结果表明,低温胁迫伤害了茄子幼苗的光合机构.     

草菇栽培菌株DNA多态性的PCR-RFLP和RAPD分析

12个草菇栽培菌株的核糖体DNA（rDNA）、线粒体DNA（mtDNA）和基因组总DNA的多态性被研究了。应用PCR－RFLP技术,扩增了rDNA5.8s+ ITS区段及mtDNA小区段,并进行限制性酶切分析,结果表明菌株间的rDNA在5.8s+ ITS区段的遗传差异小,据此只能将测试的12个菌株分为两类;而对mtDNA小区段的检测未能显示出菌株间的差异,表明测试的菌株在所研究的区段具有很高的遗传相似性。在此基础上,对基因组总DNA的RAPD分析表明,菌株两两间的遗传相似系数在0.554到0.898之间,而平均相似系数为0.707。通过平均链锁聚类方式(UPGMA)聚类,发现在相似系数0.710水平上,供试菌株可分为四大类。这些研究结果为草菇的育种提供了科学依据。     

Somatic hybridization between Solanum tuberosum and species of the S. nigrum complex: Selection of vigorously growing and flowering plants

Fusion experiments were performed between diploid (2n = 2x = 24) or tetraploid (2n = 4x = 48) potato genotypes and four species of the Solanum nigrum complex, namely S. nigrum (2n = 6x = 72), S. villosum (2n = 4x = 48), S. chenopodioides (2n = 2x = 24) or S. americanum (2n = 2x = 24 and 2n = 6x = 72). All five accessions of the S. nigrum-species were successfully hybridized with at least one of the potato genotypes. Somatic combining abilities were influenced by the ploidy level as well as the genotype of the parental species. The use of kanamycin or hygromycin resistance as cell-selectable markersystem had no influence on somatic combining ability, but such markers can be useful to improve efficient selection of somatic hybrids in sufficient numbers. At least 20% of the hybrids of each successful combination performed well in vitro. However, only 60 genotypes out of 761 somatic hybrids were vigorous as well as flowering in the greenhouse. Analysis of the DNA content of somatic hybrids could be used as a criterion for the indirect selection in vitro of hybrids that were vigorous in the greenhouse. Flowering somatic hybrids of S. nigrum (+) 2x potato and S. americanum (+) 4x potato were selected with the aim of introgression of resistance traits after recurrent backcrossing with cultivated potato. This revised version was published online in July 2006 with corrections to the Cover Date.     

两种用于鉴别山羊痘病毒和绵羊痘病毒方法的应用与比较研究

为探讨鉴别山羊痘病毒和绵羊痘病毒的分子生物学方法,对实验室分离保存的3株山羊痘病毒和2株绵羊痘病毒进行p32基因和GpCR基因扩增、克隆及序列分析.将获得的p32基因序列与GenBank上登陆的羊痘病毒p32基因进行HinfⅠ酶切位点分析表明,所有绵羊痘病毒p32基因在391 bp和691 bp处存在2处酶切位点,而山羊痘病毒仅在688 bp处存在1个酶切位点.将获得的GpCR基因序列与GenBank上登录的31条相应序列进行系统进化树分析发现,根据GpCR基因序列信息可将绵羊痘病毒和山羊痘病毒分成两个独立的进化分枝.这些结果表明,利用p32基因的HinfⅠ酶切位点信息和对GpCR基因进行分子进化分析可鉴别山羊痘病毒和绵羊痘病毒,p32基因和GpCR基因可作为鉴别绵羊痘病毒和山羊痘病毒的特异性标记基因.     

Pyramiding of genes conferring resistance to Tomato yellow leaf curl virus from different wild tomato species

Tomato (Solanum lycopersicum) production in tropical and subtropical regions of the world is limited by the endemic presence of Tomato yellow leaf curl virus (TYLCV). Breeding programmes aimed at producing TYLCV‐resistant tomato cultivars have utilized resistance sources derived from wild tomato species. So far, all reported breeding programmes have introgressed TYLCV resistance from a single wild tomato source. Here, we tested the hypothesis that pyramiding resistances from different wild tomato species might improve the degree of resistance of the domesticated tomato to TYLCV. We have crossed TYLCV‐resistant lines that originated from different wild tomato progenitors, Solanum chilense, Solanum peruvianum, Solanum pimpinellifolium, and Solanum habrochaites. The various parental resistant lines and the F₁ hybrids were inoculated in the greenhouse using viruliferous whiteflies. Control, non‐inoculated plants of the same lines and hybrids were exposed to non‐viruliferous whiteflies. Following inoculation, the plants were scored for disease symptom severity, and transplanted to the field. Resistance was assayed by comparing yield of inoculated plants to those of the control non‐inoculated plants of the same variety. Results showed that the F₁ hybrids between the resistant lines and the susceptible line suffered major yield reduction because of infection, but all hybrids were more resistant than the susceptible parent. All F₁ hybrids resulting from a cross between two resistant parents, showed a relatively high level of resistance, which in most cases was similar to that displayed by the more resistant parent. In some cases, the hybrids displayed better levels of resistance than both parents, but the differences were not statistically significant. The F₁ hybrid between a line with resistance from S. habrochaites and a line with resistance from S. peruvianum (HAB and 72‐PER), exhibited the lowest yield loss and the mildest level of symptoms. Although the resistance level of this F₁ hybrid was not statistically different from the level of resistance displayed by the 72‐PER parent itself, it was statistically better than the level of resistance displayed by the F₁ hybrids between 72‐PER and any other resistant or susceptible line.     

间种对黄曲条跳甲种群数量的影响

调查了作物之间的间种对黄曲条跳甲种群数量的影响。结果表明：白菜Brassica chinensis与葱Allium fistulosum、菜心B. parachinesis与茄子Solanum melongena间作,能显著地减轻黄曲条跳甲对白菜、菜心的为害,间作田上的黄曲条跳甲成虫的发生数量与单种田相比减少明显。在白菜地上间作上芥菜B. juncea,白菜上成虫数量逐渐减少,芥菜上的虫量逐渐增加;在芥兰B. alboglabra地上间作萝卜Raphanus sativus后,黄曲条跳甲成虫大都转移到萝卜为害,萝卜上的跳甲成虫数量高出芥兰的十倍甚至十几倍,使间作田上芥兰的黄曲条跳甲种群数量明显少于芥兰单种田,所设置的4种间作相隔距离（2m、4m、8m、15m）之间差异不显著,说明每隔约15m间种一行萝卜就能起到很好的诱集跳甲成虫效果。     

Chloroplast DNA variation in the wild potato species, Solanum acaule and S. albicans

The chloroplast DNA of Solanum acaule (109 accessions) and S. albicans (9 accessions) was investigated by restriction endonuclease analysis. Unexpectedly, all the accessions analyzed had C type chloroplast DNA in common. This suggested that S. acaule originated from a species with C type chloroplast DNA. DraI restriction digestion revealed further differentiation of C type chloroplast DNA into 8 types. The DraI polymorphism indicated the province of Salta in Argentina and the nearby regions to be a center of diversity for S. acaule. Surprisingly, S. albicans as well as S. acaule both ssp. acaule and ssp. punae, from Peru were virtually indistinguishable, although by morphology and/or cytology all three taxa are easily distinguished. This revised version was published online in July 2006 with corrections to the Cover Date.     

茄子Pn日变化及光合“午休”的生理生态子分析

以两个品种圆茄（二苠茄和快圆茄）为试材，系统研究了其光合速率（Pn）日变化及影响光合“午休”生理生态因子。结果表明，茄子晴天Pn日变化无论品种是否相同（二苠茄和快圆茄）、生育期是否相同（苗期、初花期和盛果期）、叶位是否相同（上部叶片、中部叶片和底部叶片），无一例外地表现为“双峰”曲线，即有光合“午休”现象存在，只是“午休”的时间和程度上有差异；光合“午休”时气孔导度（Gs）、蒸腾速率（Tr）、细胞间隙CO2浓度（Ci）、大气CO2浓度（Ca）均下降，而光合有效辐射（PAR）和叶片温度（TL）此时几乎处于最大值，茄子光合“午休”是由气孔因素引起的。     

High resolution map of eggplant (Solanum melongena) reveals extensive chromosome rearrangement in domesticated members of the Solanaceae

A linkage map of eggplant was constructed for an interspecific F₂ population derived from a cross between Solanum linnaeanum MM195 and S. melongena MM738. The map contains 400 AFLP^® (amplified fragment length polymorphism), 348 RFLP (restriction fragment length polymorphism) and 116 COSII (conserved ortholog set) markers. The 864 mapped markers encompass 12 linkage groups, span 1,518 cM and are spaced at an average interval of 1.8 cM. Use of orthologous markers allowed confirmation of the established syntenic relationships between eggplant and tomato chromosomes and helped delineate the nature of the 33 chromosomal rearrangements and 11 transpositions distinguishing the two species. This genetic map provides a 2- to 3-fold improvement in marker density compared to previously published interspecific maps. Because the interspecific mapping population is rich in morphological variation, this greater genome saturation will be useful for QTL (quantitative trait locus) analyses. The recent release of the tomato genome sequence will provide additional opportunities for exploiting this map for comparative genomics and crop improvement.     

Intra- and interspecific transmission of androgenetic competence in diploid potato species

Summary In an attempt to determine the transmission of androgenetic competence, 10 families resulting from intra- and interspecific hybrids including three reciprocal hybrids were examined in anther culture. Hybrid families were generated between competent clones of Solanum phureja and incompetent clones of S. phureja, S. microdontum and S. berthaultii. S. phureja clones PP5 and A95 (derived by androgenesis of a 2n microspore of PP5) were found to be consistently competent for androgenesis. Androgenetic competence was observed to segregate in all hybrid families with some highly responsive and some unresponsive genotypes in all families. A total of 9,465 cultured anthers have yielded 936 embryoids and 91 plants, including 29 monoploids. The cytoplasm of species lacking competence appeared to have greater influence on the expression of androgenesis in intraspecific than in interspecific hybrids. Expression of androgenesis varied among half-sib hybrid families indicating that competence for androgenesis was influenced by the parents lacking competence. The anther culture data on a backcross between a highly responsive hybrid and its unresponsive parent indicated that competence may be under control of a single dominant gene.     

Segregation of isozymes in selfed progenies of a synthetic amphidiploid between Solanum integrifolium and S. melongena

Isozyme and cytogenetic analyses were performed on selfed progenies of a synthetic amphidiploid between scarlet eggplant, Solanum integrifolium (= S. aethiopicum),and eggplant, Solanum melongena `DMP', for estimating genetic uniformity. Isozymes in the 379 examined seedlings segregated into five genotypes (phenotypes) each at the four loci examined, Pgd-2 of phosphogluconate dehydrogenase (E.C.1.1.1.43), Idh-2 of isocitrate dehydrogenase (E.C.1.1.1.41), Pgm-2 of phosphoglucomutase (E.C.2.7.5.1)and Skdh-1 of shikimate dehydrogenase (E.C.1.1.1.25), indicating that the selfed seedlings were not genetically uniform. Most of the examined 15 selfed seedlings exhibited a somatic chromosome number of 48, that is the same number of the synthetic amphidiploid, whereas isozyme genotypes among them were variable. It is suggested that the segregation of isozymes was not caused by variation of chromosome number but by genetic segregation of isozyme genes. The genome of the synthetic amphidiploid was indicated to be unstable. This revised version was published online in July 2006 with corrections to the Cover Date.     

Initial development of a set of introgression lines from Solanum peruvianum PI 126944 into tomato: exploitation of resistance to viruses

Resistance to Tomato yellow leaf curl virus (TYLCV) and Tomato spotted wilt virus (TSWV), among other diseases, has been reported in Solanum peruvianum PI 126944. Introgression lines (ILs) from S. peruvianum PI 126944 into the genetic background of cultivated tomato (S. lycopersicum) are being developed. Several generations were derived from three interspecific hybrids previously obtained. A lot of crosses and embryo rescue were required until the third backcross, due to the high degree of incompatibility existing between tomato and PI 126944. Crosses between F₁ plants were made to obtain a pseudo-F₂ generation. The same procedure was followed up to the pseudo-F₆ generation. Additional crosses between plants of different generations were made in order to increase progeny. Of 263 molecular markers tested, 105 were polymorphic between tomato and PI 126944. This set of polymorphic markers consisted of 90 simple sequence repeats (SSR) and 15 cleaved amplified polymorphic sequences (CAPS). The amount of the S. peruvianum genome was reduced in advancing generations and this was coupled in some cases with a reduction of incompatibility. However, the S. peruvianum genome was almost completely represented among the different plants of the most advanced generations. ILs will be basically developed from them. Some of the generations developed were resistant to TYLCV and TSWV.     

猪传染性胃肠炎病毒S基因A抗原位点的克隆及原核表达载体的构建

【研究目的】对猪传染性胃肠炎病毒S基因A抗原位点进行克隆和原核表达载体的构建。【方法】参考GenBank上公布的TGEV S基因A抗原位点序列,应用Primer6.0设计一对含酶切位点的引物,用于RT-PCR扩增A抗原位点目的片段,将扩增产物连接于paesy-T克隆载体上构建克隆载体,用EcoR I和Xhol I对表达载体PET-32a（+）和重组质粒进行酶切,将酶切产物亚克隆至PET-32a（+）多克隆位点上,连接、转化至BL21(DE3),构建A位点的原核表达载体,并对阳性重组质粒进行酶切、PCR和测序鉴定。【结果】所扩增的目的片段的大小为534bp,与原核表达载体连接后,经核苷酸及推导的氨基酸序列分析表明,该基因与其它猪传染性胃肠炎病毒相应基因具有很高的同源性,说明成功地构建了TGEV S 基因A抗原位点的原核表达载体。【结论】TGEV S 基因A抗原位点原核表达载体的成功构建,填补了中国国内单独针对此位点进行研究的空白,也为TGEV诊断方法的建立提供良好的技术基础。     

Identification and characterization of microsatellites in eggplant

The potential of microsatellite markers for use in genetic studies in eggplant, Solanum melongena, has been evaluated. A genomic library of eggplant was screened for GA and GT repeat motifs to isolate microsatellite clones. The frequency of each repeat motif in the eggplant genome was found to be every 3200 kb for GA repeats and every 820 kb for GT repeats. Sixty‐one per cent of GT repeats were found to directly flank AT repeats. A total of 37 polymerase chain reaction (PCR) primer pairs were designed, 23 of which amplified a single product or several products. The level of microsatellite polymorphism was evaluated by using S. melongena lines and related Solanum species. Two to six alleles per primer pair were displayed in the S. melongena lines and two to 13 alleles were displayed in the Solanum relatives. Seven microsatellites showed polymorphism between parental lines of the mapping population and segregated in a codominant Mendelian manner. These microsatellite loci were distributed throughout the linkage map.     

Production of somatic hybrid plants between Iris ensata Thunb. and I. germanica

Wide hybridization that cannot be attained through conventional sexual crosses, can now be approached by somatic hybridization. Protoplasts of I. ensata and I. germanica were fused by electrofusion. For the selection of somatic hybrids, protoplasts of I. ensata which did not form colonies in protoplast culture and protoplasts of I. germanica which had regeneration ability for only albino shoot were used in symmetric fusion. On the other hand, the protoplasts of I. ensata and I. germanica protoplasts which were inactivated by iodoacetamide (IOA) treatment were used in asymmetric fusion. Five-six months after cell fusion, green plants were obtained in the symmetric and asymmetric fusion. In the random amplified polymorphic DNA (RAPD) analysis, the green plants had bands specific to both parental species. Therefore, these plants were somatic hybrids between I. ensata and I. germanica. This revised version was published online in July 2006 with corrections to the Cover Date.     

Organelle based molecular analyses of the genetic relatedness of cultivated alfalfa (Medicago sativa L) to Medicago edgeworthii Sirjaev, and Medicago ruthenica (L.) Ledebour

Medicago edgeworthii Sirjaev and M. ruthenica (L.) Ledebour are allogamous, diploid (2n = 2x = 16) perennials with flat pods.Medicago edgeworthii is indigenous to the Himalayas and alpine areas west to Afghanistan, and Medicago ruthenica is found in Siberia, Mongolia, and Manchuria on open hillsides and mixed grass steppes. Because both species have a remarkable ability to survive extreme cold and poor soils, the possibility of hybridizing them with alfalfa (M. sativa L.) is being investigated. The objective of this research was to conduct an organelle based molecular assessment of the genetic relatedness of cultivated alfalfa (2n = 4x = 32) to M. edgeworthii and M. ruthenica. A hypervariable, intergenic region of cpDNA was amplified, and mtDNA was amplified with two primer pairs developed from soybean (Glycine max L.) mtDNA sequences. Mean Nei and Li genetic distances (GDs) between alfalfa and M. edgeworthii and alfalfa and M. ruthenica were 0.56 and 0.48 (mtDNA), and 0.33 and 0.30 (cpDNA), respectively. Intra specific GDs were 0.37 (mtDNA) and 0.25 (cpDNA) for M. edgeworthii; 0.42 (mtDNA) and 0.15 (cpDNA) for M. ruthenica; and 0 = 0.50 (mtDNA) and 0 = 0.23 (cpDNA) for alfalfa. Cluster analyses grouped someM. edgeworthii and M. ruthenica entries with alfalfa entries. There is some chance that alfalfa and M. edgeworthii entries which clustered closely could be hybridized; chances of alfalfa × M. ruthenica hybridizations appear to be more problematic. This revised version was published online in July 2006 with corrections to the Cover Date.