An allele of dihydroflavonol 4-reductase associated with the ability to produce red anthocyanin pigments in potato (Solanum tuberosum L.)

The potato R locus is necessary for the production of red pelargonidin-based anthocyanin pigments in any tissue of the plant, including tuber skin and flower petals. The production of pelargonidins in plants requires the activity of dihydroflavonol 4-reductase (DFR) to catalyze the reduction of dihydrokaempferol into leucopelargonidin. To test the hypothesis that potato R encodes DFR, portions of both dfr alleles were sequenced from a diploid potato clone known to be heterozygous Rr. Sequence comparison revealed a polymorphic BamHI restriction site. The presence or absence of this site was monitored in three diploid populations that segregated for R, as well as in a wide range of tetraploid breeding clones and cultivars, by amplifying a fragment of dfr and digesting the products with BamHI. An identically sized dfr restriction fragment lacking the BamHI site was present in all potato clones that produced red anthocyanin pigments, while the same fragment was absent in many potato clones with white tuber skin and flowers. An independent RFLP test using DraI to detect sequence polymorphism was performed on a subset of the potato clones. This test also revealed dfr-derived bands that were present in all red-colored potatoes and absent in several white clones. The presence of shared restriction fragments in all red-colored potatoes provides strong evidence that R does indeed code for DFR. The data are also consistent with a 48 year-old hypothesis by Dodds and Long, that R was selected just once during the domestication of potato. A cDNA clone corresponding to the red allele of dfr was sequenced and compared to two other alleles. The red allele is predicted to encode a 382 amino acid protein that differs at ten amino acid positions from the gene products of the two alternative alleles. Several of these differences map in a region known to influence DFR substrate specificity in Gerbera.Communicated by J. Dvorak     

The potato P locus codes for flavonoid 3′,5′-hydroxylase

The potato P locus is required for the production of blue/purple anthocyanin pigments in any tissue of the potato plant such as tubers, flowers, or stems. We have previously reported, based on RFLP mapping in tomato, that the gene coding for the anthocyanin biosynthetic enzyme flavonoid 3,5-hydroxylase (f35h) maps to the same region of the tomato genome as P maps in potato. To further evaluate this association a Petunia f35h gene was used to screen a potato cDNA library prepared from purple-colored flowers and stems. Six positively hybridizing cDNA clones were sequenced and all appeared to be derived from a single gene that shares 85% sequence identity at the amino acid level with Petunia f35h. The potato gene cosegregated with purple tuber color in a diploid F₁ sub-population of 37 purple and 25 red individuals and was found to be expressed in tuber skin only in the presence of the anthocyanin regulatory locus I. A potato f35h cDNA clone was placed under the control of a doubled CaMV 35S promoter and introduced into the red-skinned cultivar Désirée. Tuber and stem tissues that are colored red in Désirée were purple in nine of 17 independently transformed lines.An erratum to this article can be found at     

A fluorogenic 5′ nuclease (TaqMan) assay to assess dosage of a marker tightly linked to red skin color in autotetraploid potato

We have recently identified an allele of dihydroflavonol 4-reductase (dfr) that cosegregates with the ability of potato (Solanum tuberosum L) to produce red pelargonidin-based anthocyanin pigments. A rapid assay to assess dosage of this allele in cultivated potato, an autotetraploid, would be useful for breeding programs that develop red-skinned cultivars. To identify regions of dfr that are conserved between alleles, as well as regions that are variable, a portion of the gene was sequenced from several cultivated and wild potato clones. In one region the sequence of the 'red' dfr allele differed at two nucleotide positions from the three other sequence classes observed. A fluorogenic oligonucleotide probe labeled with 6-FAM was designed to anneal specifically to the red allele in this region, while a second probe labeled with VIC was designed to anneal to the 'not-red' dfr alleles. PCR primers that annealed to conserved sequences flanking the variable region were also developed. When subjected to a fluorogenic 5 nuclease (TaqMan) allelic discrimination assay all diploid clones tested clustered into three distinct groups based on the relative amounts of FAM and VIC released. These three groups represented clones homozygous for the red allele, heterozygous for the red allele, and homozygous for the not-red allele(s). When tetraploid clones were tested they separated into five distinct clusters, three of which were shared with diploid clones. The five clusters were interpreted to represent clones quadruplex, triplex, duplex, simplex and nulliplex for the red dfr allele. This interpretation was supported by monitoring the segregation of red allele dosage in several tetraploid crosses. To the best of our knowledge this is the first report of a fluorogenic 5 nuclease assay being used for allelic discrimination in an autopolyploid.Communicated by J.W. Snape     

三七块根紫色素的花色苷本质及其含量和总皂苷含量的正相关性

三七是中国云南省的"第一药材",云南文山三七是三七的道地药材。三七块根的横截面为黄白色至紫色。紫色块根约占研究块根总数的28 .21 %,其中柱鞘、内皮层、皮层或表皮为紫色。特征颜色反应和紫外——可见光谱表明:三七块根紫色素属于黄酮类化合物,可能含有酚性邻位二羟基,不含类胡萝卜素、查耳酮、噢哢、异黄酮、儿茶素。花色苷和/或其苷元花色素奠定了紫色块根着色的基础,其他的非红色的黄酮类化合物起共色素的作用。块根的平均花色苷含量和平均总皂苷含量均以纯紫色块根的为最高,其次是黄紫混合色块根的,纯黄色的最低。块根的花色苷含量差异达到极显著水平,但总皂苷含量差异却没有达到显著水平。每个块根都含有不同量的花色苷,随花色苷量的增加,块根的紫色一般逐渐明显。块根的花色苷含量与其总皂苷含量之间呈显著正相关,相关系数r=0 .355。本文可为三七块根颜色呈现的机理探索及其色素的分子结构鉴定提供参考。     

Protective effect of supplemental anthocyanins on Arabidopsis leaves under high light

Ten anthocyanin components have been detected in roots of purple sweet potato (Ipomoea batatas Lam.) by high‐performance liquid chromatography coupled to diode array detection and electrospray ionization tandem mass spectrometry. All the anthocyanins were exclusively cyanidins or peonidin 3‐sophoroside‐5‐glucosides and their acylated derivatives. The total anthocyanin content in purple sweet potato powder obtained by solid‐phase extraction was 66 mg g^?1. A strong capacity of purple sweet potato anthocyanins (PSPA) to scavenge reactive oxygen species (superoxide, hydroxyl radical) and the stable 1,1‐diphenyl‐2‐picrylhydrazyl organic free radical was found in vitro using the electron spin resonance technique. To determine the functional roles of anthocyanins in leaves in vivo, for the first time, supplemental anthocyanins were infiltrated into leaves of Arabidopsis thaliana double mutant of the ecotype Landsberg erecta (tt3tt4) deficient in anthocyanin biosynthesis. Chlorophyll fluorescence imaging showed that anthocyanins significantly ameliorated the inactivation of photosystems II during prolonged high‐light (1300 µmol m^?2 s^?1) exposure. Comet assay of DNA revealed an obvious role of supplemental PSPA in alleviating DNA damage by high light in leaves. Our results suggest that anthocyanins could function in vitro and in vivo to alleviate the direct or indirect oxidative damage of the photosynthetic apparatus and DNA in plants caused by high‐light stress.     

Aspects of tuber resistance in hybrid potatoes to potato tuber worm

Tubers produced from crosses between the wild potato, Solanum berthaultii Hawkes (Solanaceae), and the cultivated species Solanum tuberosum L. (Solanaceae) are resistant to potato tuber worm (PTW), Phthorimaea operculella (Zeller) (Lepidoptera: Gelechiidae), infestation compared to those of the popular commercial North American cultivars Allegany, Atlantic, Chieftain, Katahdin, MaineChip, NorDonna, Norwis, Russet Norkotah, Snowden, and Yukon Gold. Given a choice between Atlantic and hybrid tubers, female PTW deposited ca. 50% fewer eggs on hybrid tubers than on those of Atlantic; larval survival and production of prepupae on hybrid tubers were reduced similarly. Time needed for neonates to penetrate eye buds was ca. 100 min greater on hybrid tubers compared to that on cv. Atlantic. Periderm of hybrid tubers is thicker than that of cv. Atlantic and may contribute to the delay in larval penetration of tubers and the success of initial establishment.     

菊花不同花色品种中花青素苷代谢分析

应用高效液相色谱和多级质谱联用技术(HPLC-ESI-MSⁿ), 分析菊花(Chrysanthemum × morifolium)白色、粉色、红色、紫色、红紫色和墨色6个色系共计82个品种中花青素苷合成过程的中间产物和最终产物, 发现从白色、粉色、红色、紫色、红紫色到墨色花青素苷含量快速增加, 分别为4.68、111.60、366.89、543.56、1 220.36和2 674.95 μg·g^–1, 不同色系间花青素苷的含量差异显著(P<0.01), 花青素苷含量越高花色越深; 墨色菊花品种中总类黄酮含量显著高于其它花色品种(P<0.01), 其它不同色系间总类黄酮含量差异不显著(P>0.05); 随着菊花花色变深, 从柚皮素分支到圣草酚的代谢流, 以及从圣草酚分支到矢车菊素苷的代谢流比例增加。花青素苷成分分析发现: 菊花中只含有矢车菊素苷类化合物。根据花青素苷代谢成分分析结果绘制了菊花中花青素苷代谢路径图, 即在菊花类黄酮代谢途径中只存在矢车菊素苷代谢分支途径;菊花不同色系在柚皮素和圣草酚2个关键代谢分支点上向不同方向代谢流的分配比例不同, 造成花青素苷产物含量不同,导致不同花色。以上研究结果为菊花花色改良的分子育种提供了理论依据。     

Differential gene expression analysis of ‘Granny Smith’ apple (Malus domestica Borkh.) during fruit skin coloration

‘Granny Smith’ apples growing under normal sunlight develop green skin, whereas the peel turns red due to anthocyanin accumulation after the removal of a bagging treatment. Two anthocyanins, Cyanidin 3-O-galactoside (cy3-gal) and Cyanidin 3-O-arabinoside (cy3-ara), were detected in the red ‘Granny Smith’ apple peels, and cy3-gal was determined to be chiefly responsible for the red color. The content of cy3-gal was more than 98% of the total anthocyanin in the red ‘Granny Smith’ peels. To better understand the molecular basis of anthocyanin biosynthesis in ‘Granny Smith’ apples, we performed a quantitative real-time PCR (qRT-PCR) analysis of anthocyanin biosynthetic genes (MdCHS, MdF3H, MdDFR, MdANS, MdUFGT, and MdMYB1). Our results indicate that the expression of these genes (except MdCHS) was associated with increased anthocyanin accumulation in the skin of ‘Granny Smith’ apples. Four selected genes obtained from the ‘Granny Smith’ skin cDNA library, phytoene synthase (PSY), WD40 repeat protein, polygalacturonase (PG), and galactosidase (GAL), were also confirmed by qRT-PCR. We found that these genes were differently expressed during ‘Granny Smith’ apple skin coloration, suggesting that they are directly or indirectly involved in pigment accumulation. In conclusion, anthocyanin biosynthesis in ‘Granny Smith’ apples is the result of interactions between multiple enzymes in the anthocyanin biosynthesis pathway, and the coloring mechanism of ‘Granny Smith’ apples may be similar to that of red-skinned cultivars.     

The effect of soil- and tuber-borne inoculum on the incidence of potato gangrene

Under optimum growing conditions neither tuber- nor soil-borne Phoma exigua var. foveata inoculum appreciably affected stand or yield of the subsequent potato crop. Seed tubers with gangrene rots caused high levels of stem and tuber symptoms when planted in var. foveata contaminated or uncontaminated land; contaminated seed tubers with no rots also produced progeny with a high gangrene potential. Sufficient soil-borne inoculum was carried over in land that produced a gangrene affected crop in the previous year to override the effect of tuber disinfection. Effective gangrene control was achieved by a combination of tuber disinfection shortly after harvest over successive years with a 1 in 5 yr potato crop rotation. Gangrene rots usually developed through injuries to the tuber periderm, rots in other tubers being associated with pustules of powdery scab (Spon-gospora subterranea).     

THE DIFFERENTIATION OF PIGMENTATION IN FLOWER PARTS. III. METABOLISM OF SOME EXOGENOUS ANTHOCYANINS BY DETACHED PETALS OF IMPATIENS BALSAMINA

The anthocyanins of mature petals of Impatiens balsamina L. are distinct from the pigments found in vegetative tissue. In the red genotype (llHHP^rP^r) a sequential elaboration of the characteristic anthocyanins has been previously demonstrated through the examination of buds at successive stages of development. The metabolism of anthocyanins, especially pelargonidin-3-mono-glucoside, was examined by infiltration into developing petals of a genetically white strain. This anthocyanin appears to play a central role in the biochemical sequences involved and it has been observed that the genetically white flowers possess the enzymatic potential to metabolize this substrate, producing the same final products which are produced in the red genotype. There is a pattern of change in the relative amounts of each anthocyanin during the incubation period which follows closely the pattern which occurs during normal development of the colored genotypes. This indicates that the enzymes which are normally produced in the colored flowers are also produced in flowers which never produce anthocyanins. The metabolic capabilities of several other genetic strains and the influence of light and puromycin have been examined.     

铁筷子花瓣中花青素苷成分及含量对其呈色的影响北大核心CSCD

该研究以7个品种铁筷子(Helleborus thibetanus Franch.)为试验材料,借助目视测色、RHSCC比色卡、色差仪进行花色表型的测定,采用高效液相色谱法-光电二极管阵列检测方法(HPLC-DAD)及高效液相色谱-电喷雾离子化-质谱联用技术(HPLC-ESI-MS)测定分析铁筷子花瓣中花青素苷成分及含量,以探究不同品种铁筷子的花色与花青素苷成分及含量之间的关系。结果显示:(1)紫色系品种花瓣的a*值最高b*值最低,黄色系品种花瓣的b*值最高a*值最低,不同品种的铁筷子花色越深L*值越低。(2)从5个有花青素苷积累的铁筷子品种中检测出11种花青素苷成分,分别为6种矢车菊素苷,4种飞燕草素苷,1种矮牵牛素苷;供试的铁筷子材料中红色系2个品种的花青素苷含量最高,紫色系品种次之;矢车菊素苷与飞燕草素苷为影响铁筷子花瓣呈色的主要色素物质。(3)不同种类的花青素和修饰基团的差异,导致铁筷子花瓣呈现不同的色彩,含有多种酰基化修饰的飞燕草素苷使铁筷子花色蓝移进而使花色加深。(4)相关分析表明,铁筷子花瓣的L*值与a*值呈显著负相关关系,与b*值呈显著的正相关关系;L*值与总花青素苷含量呈显著负相关关系,且随着花青素苷含量的累积a*值增加,花色红移。研究表明,花青素苷的成分及含量是导致铁筷子花瓣呈现不同颜色的主要原因,矢车菊素苷和飞燕草素苷的互作以及酰基化的修饰使铁筷子呈现不同程度的紫色,花青素苷的不同累积量影响了花瓣颜色的明暗变化,从而使铁筷子花瓣颜色丰富。     

Increased accumulation of anthocyanins in transgenic potato tubers by overexpressing the 3GT gene

In order to explore a biotechnological method for improving potato tuber color and creating plants with increased anthocyanin contents, a potato UDP-glucose: flavonoid-3-O-glucosyltransferase (3GT) gene was inserted behind the GBSSI promoter of pBin19, and this construct was introduced into Solanum tuberosum L. cultivar Désirée plants by Agrobacterium-mediated transformation. Six independent transgenic lines overexpressing the 3GT gene were identified by PCR and Southern blot analysis from 18 kanamycin-resistant plants. Due to the expression of 3GT gene, the tuber color and the anthocyanin content were enhanced noticeably in the transgenic plants compared to the wild-type control plants. This result suggests that the 3GT gene can potentially be used to improve potato color and enhance levels of antioxidants in the diet.