Molecular analysis of two Chinese pear (Pyrus bretschneideri Rehd.) spontaneous self-compatible mutants, Yan Zhuang and Jin Zhui

Yan Zhuang and Jin Zhui are spontaneous bud mutants of Chinese pear ( Pyrus bretschneideri Rehd.) from Ya Li. Both fruit set rate and seed number after self-pollination, together with pollen tube growth, prove that Yan Zhuang and Jin Zhui are self-compatible. The fruit set rate and seed number after cross-pollination suggest that the self-compatibility of Yan Zhuang and Jin Zhui may be due to natural mutations of the stylar S allele and pollen S allele, respectively. PCR amplification of the S-RNase gene in self-pollinated progeny of Yan Zhuang and Jin Zhui show that they contain point mutations in the stylar S₂₁ allele and pollen S₃₄ allele, respectively. The cDNA sequence of the Yan Zhuang stylar S-RNase gene revealed that the 182nd nucleotide of the S₂₁-RNase (cDNA) sequence had been substituted resulting in a Gly to Val mutation, and this might affect the stability of the S-RNase. In addition, Western blotting showed that one Yan Zhuang stylar S-RNase was absent and the expression level of another S-RNase protein was decreased compared to Ya Li. Therefore, we suggest that the self-compatibility of Yan Zhuang is caused by a point mutation in an S₂₁-RNase nucleotide.     

Genome-wide identification and comparative analysis of the ADH gene family in Chinese white pear (Pyrus bretschneideri) and other Rosaceae species

Alcohol dehydrogenase (ADH) is essential to the formation of aromatic compounds in fruits. However, the evolutionary history and characteristics of ADH gene expression remain largely unclear in Rosaceae fruit species. In this study, 464 ADH genes were identified in eight Rosaceae fruit species, 68 of the genes were from pear and which were classified into four subgroups. Frequent single gene duplication events were found to have contributed to the formation of ADH gene clusters and the expansion of the ADH gene family in these eight Rosaceae species. Purifying selection was the major force in ADH gene evolution. The younger genes derived from tandem and proximal duplications had evolved faster than those derived from other types of duplication. RNA-Seq and qRT-PCR analysis revealed that the expression levels of three ADH genes were closely correlated with the content of aromatic compounds detected during fruit development.     

'砀山酥梨'的组织培养和脱毒快速繁殖

1植物名称白梨品种‘砀山酥梨’（Pyrus bretschneideri Rehd．）。 2材料类别茎尖。 3培养条件分化培养基：（1）改良MS＋6-BA2．0mg．L^-1（单位下同）＋NAA0．5＋0．5％活性炭（AC）;丛生芽诱导培养基：（2）MS＋NAA0．5＋6．BA0．5、1．0、1．5、2．0、3．0;增殖继代培养基：（3）MS＋6．BA1．0＋GA31．0＋NAA0．2、0．5、1．0、1．5、2．0;生根培养基：（4）ASH＋间苯三酚（PG）80＋IBA1．0＋NAA0．5,（5）ASH＋IBA1．0＋NAA0．5。     

Identification and characterization of invertase family genes reveal their roles in vacuolar sucrose metabolism during Pyrus bretschneideri Rehd. fruit development

23 invertase (PbrInvs) genes, including eight vacuolar invertases (PbrvacInvs), five cell wall invertases (PbrcwInvs) and 10 alkaline/neutral invertases (PbrA/N-Invs), were identified from P. bretschneideri Rehd. genome, with diverse chromosome locations, cis-acting elements, gene structures and motifs. Their expression profiles were tissue-specific, and postharvest light or temperature treatment would alter their expression profiles. During ‘Dangshansuli’ pear development, in association with visual/inner quality change was the alternations of invertase activity and the expression profiles of PbrInvs. In combination with results of subcellular sugar distribution as well as correlation analysis among sugar content, invertase activity and PbrInv mRNA abundance, PbrvacInv1 might be involved in sucrose decomposition during pear development. PbrvacInv1-GFP fusion protein mainly accumulated on the tonoplast (vacuolar membrane); meanwhile, transient overexpression of PbrvacInv1 in pear fruit would upregulate vacInv activity, causing higher fructose and lower sucrose when compared with that of the control. Furthermore, invertase inhibitor 5 (PbrInvInh5) could interact with PbrvacInv1.     

Phylogenetic and Expression Analysis of Mn-CDF Transporters in Pear (Pyrus bretschneideri Rehd.)

Plant Molecular Biology Reporter - Metal tolerance proteins (MTPs) play important roles in heavy metal homeostasis and tolerance in plants. As a part of the MTP family, manganese-cation diffusion...     

砀山酥梨4CL基因家族的全基因组鉴定与分析

4-香豆酸：辅酶A连接酶(4CL)基因是植物调控木质素代谢、参与类黄酮和其他次生代谢产物合成的关键基因之一,而木质素的合成及聚合在细胞壁沉积导致部分薄壁细胞次生壁加厚形成石细胞。为更好了解砀山酥梨中4CL基因的种类和数量,本文利用砀山酥梨基因组的氨基酸和cDNA数据库对4CL基因家族进行筛选,分析了砀山酥梨基因组中4CL基因的种类、进化关系、物理定位、以及基因结构和保守基序。结果显示在砀山酥梨基因组中发现并初步确定了29个4CL基因,通过基因的定位分析发现除了4、8、11、12号染色体上没有4CL基因之外,其他染色体上都存在4CL基因;并且在9、17号染色体上还存在着基因簇。通过基因结构和进化树之间的比较,进一步确定了基因结构和进化之间的相互联系。研究结果为砀山酥梨4CL基因功能的深入分析奠定了基础。     

Cloning of cDNAs encoding cell-wall hydrolases from pear (Pyrus communis) fruit and their involvement in fruit softening and development of melting texture

'La France' pear ( Pyrus communis L.) fruit stored at 1°C for 1 month (short-term storage) before transfer to 20°C softened and developed a melting texture during ripening, whereas fruit stored for 5 months (long-term storage) before transfer to 20°C softened but did not develop a melting texture. To clarify the mechanisms involved in fruit softening and textural changes, the cDNAs encoding cell-wall hydrolases were isolated by RT-PCR, and their expression and localization were investigated in 'La France' pears. Genes encoding three polygalacturonases (PG; EC 3.2.1.15), four pectin methylesterases (PME; EC 3.1.1.11), one α -arabinofuranosidase (ARF; EC 3.2.1.55), three β -galactosidases (GAL; EC 3.2.1.23), and two endo-1,4- β - d -glucanases (Cel; EC 3.2.1.4) were isolated. Among these 13 isolated genes, PcPG1 was the only gene for which the mRNA expression levels increased in both the short- and long-term stored fruits. This suggested that PcPG1 is involved in fruit softening rather than in the development of the melting texture. In contrast, the expression levels of PcPG3 , PcPME1 , PcPME2 , PcPME3 , PcGAL1 , PcGAL2 , and PcCel2 increased during ripening only in the short-term stored fruit. These genes might thus be involved in the development of the melting texture.     

中国梨2个自交不亲和新等位基因(S等位基因)的分子鉴定

自交不亲和是显花植物的一种重要生殖生理现象,为探明中国梨的自交不亲和特性,对‘锦香’(Pyrus bretschneideri cv. Jinxiang)和‘鹅酥’(Pyrus bretschneideri cv. Esu)2个中国梨品种进行了基因组PCR特异扩增、S基因序列分析及田间杂交授粉试验。结果确定它们各含1个新S-RNA酶基因,分别命名为S37-和S38-RNase,GenBank序列号为DQ839238和DQ839239。生物信息学分析结果表明,S37-和S38-RNA酶的推导氨基酸序列与S1-至S36-RNA酶36个梨S基因具有相同的、高度保守的C1和C2区,但其高变区与S1-至S36-RNA酶差异较大,其中与S15的差异最小,只有3个氨基酸不同。在推导的氨基酸水平上,S37与S38有96%的序列相似性,但两者与S15的相似性更高,皆为98%,与S32的相似性最低,都只有63%;S37和S38的内含子较大,分别为786bp和723bp,与S15的777bp大小接近。最后,经分析验证确定‘锦香’和‘鹅酥’的S基因型分别为S34S37和S15S38。     

Partitioning of (13)C-photosynthate from spur leaves during fruit growth of three Japanese pear (Pyrus pyrifolia) cultivars differing in maturation date

BACKGROUND AND AIMS: In fruit crops, fruit size at harvest is an important aspect of quality. With Japanese pears (Pyrus pyrifolia), later maturing cultivars usually have larger fruits than earlier maturing cultivars. It is considered that the supply of photosynthate during fruit development is a critical determinant of size. To assess the interaction of assimilate supply and early/late maturity of cultivars and its effect on final fruit size, the pattern of carbon assimilate partitioning from spur leaves (source) to fruit and other organs (sinks) during fruit growth was investigated using three genotypes differing in maturation date. METHODS: Partitioning of photosynthate from spur leaves during fruit growth was investigated by exposure of spurs to (13)CO(2) and measurement of the change in (13)C abundance in dry matter with time. Leaf number and leaf area per spur, fresh fruit weight, cell number and cell size of the mesocarp were measured and used to model the development of the spur leaf and fruit. KEY RESULTS: Compared with the earlier-maturing cultivars 'Shinsui' and 'Kousui', the larger-fruited, later-maturing cultivar 'Shinsetsu' had a greater total leaf area per spur, greater source strength (source weight x source specific activity), with more (13)C assimilated per spur and allocated to fruit, smaller loss of (13)C in respiration and export over the season, and longer duration of cell division and enlargement. Histology shows that cultivar differences in final fruit size were mainly attributable to the number of cells in the mesocarp. CONCLUSIONS: Assimilate availability during the period of cell division was crucial for early fruit growth and closely correlated with final fruit size. Early fruit growth of the earlier-maturing cultivars, but not the later-maturing ones, was severely restrained by assimilate supply rather than by sink limitation.     

莱阳茌梨果实褐变与多酚氧化酶及酚类物质区域化分布的关系

在气调贮藏中,莱阳在梨果心的组织褐变在时间上先于果肉,在程度上重于果肉。在果肉内中部3/5易褐变。褐变部位的不均一性与酚类物质在果实中分布的不均一密切相关,酚类物质含量越高,褐变越严重。层析法鉴定出绿原酸、桂皮酸、阿魏酸和咖啡酸,其中以绿原酸为多酚氧化酶的最适底物。酶法分离纯化了原生质体和液泡,表明酚类物质存在于液泡内,多酚氧化酶在细胞质中。     

Characterization of the pectin methyl-esterase gene family and its function in controlling pollen tube growth in pear (Pyrus bretschneideri)

Pectin methyl-esterases (PMEs) play crucial roles in plant growth. In this study, we identified 81 PbrPMEs in pear. Whole-genome duplication and purifying selection drove the evolution of PbrPME gene family. The expression of 47 PbrPMEs was detected in pear pollen tube, which were assigned to 13 clusters by an expression tendency analysis. One of the 13 clusters presented opposite expression trends towards the changes of methyl-esterified pectins at the apical cell wall. PbrPMEs were localized in the cytoplasm and plasma membrane. Repression of PbrPME11, PbrPME44, and PbrPME59 resulted in the inhibition of pear pollen tube growth and abnormal deposition of methyl-esterified pectins at pollen tube tip. Pharmacological analysis confirmed that reduced PbrPME activities repressed the pollen tube growth. Overall, we have explored the evolutionary characteristics of PbrPME gene family and found the key PbrPME genes that control the growth of pollen tube, which deepened the understanding of pear fertility regulation.     

鸭梨PbHCT3基因的克隆及表达分析

羟基肉桂酰CoA:莽草酸/奎宁酸羟基肉桂酰转移酶(HCT)是植物绿原酸合成的关键酶之一.该研究以鸭梨为材料,采用同源基因克隆的方法,克隆出一个鸭梨的HCT基因,命名为PbHCT3.PbHCT3基因cDNA全长序列为1 731bp,基因编码序列长度为1 317bp,编码438个氨基酸,含有酰基转移酶的2个保守序列HHAAD和DFGWG及保守结构域MVVNVTVRES.实时荧光定量PCR分析表明:PbHCT3基因在幼果果皮、果肉、果心、幼叶及花蕾期花瓣中的表达量较高.随着鸭梨果实生长,果实中PbHCT3基因表达量逐渐降低.研究表明,PbHCT3基因与鸭梨幼嫩组织的生长发育有密切关系.     

Occurrence and Latent Infection of Alternaria Rot of Pingguoli Pear (Pyrus bretschneideri Rehd. cv. Pingguoli) Fruits in Gansu, China

Alternaria rot of Pingguoli pear occurred after latent infection. Fruit surfaces were asymptomatic within 60 days storage under cold condition (0°C, RH 85–90%), but black‐grey hyphae could be seen in the lenticels or calyx tube of Pingguoli pear after 90 days of storage. The tissue collapsed and resulted in visible black spots as the hyphae spread over the fruit. Average incidence of Alternaria rot of fruits from an orchard in Gansu was 28.86% at 100 days of storage. The main fungus isolated from the Alternaria rot on stored Pingguoli pear was identified as Alternaria alternata (Fr. : Fr.) Keissl. This pathogen was able to initially infect the fruit via two pathways during the growing season, and then remain in a latent state. The fungus first colonized the styles at the full‐blossoming stage, and then grew into the carpel cavities progressively after 50 days from petal fall. The percentage latent infection of A. alternata was up to 45% in the carpel cavity until the harvest time. The fungus also attacked fruit surfaces and remained latent in the fruit peel during fruit development. The percentage of A. alternata latent infection at the calyx end, middle part and stem end of the fruit peel was 40%, 24% and 42.8%, respectively, at harvest time.     

河北鸭梨病果上链格孢分离系的形态及分子特性研究

A total of 123 Alternaria isolates were obtained from roting fruits of Pyrus bretschneideri "Ya Li" collected in Hebei Province, China. The isolates, according to morphological characteristics of conidia and sporulation patterns, were segregated into four groups: A. alternata group, A. tenuissima group, A. yaliinficiens group and Alternaria sp. group. Molecular characteristics of part of the isolates were determined using random amplified polymorphism DNA (RAPD) analysis. Based on cluster analysis of RAPD data, large-and small-spored Alternaria spp. were evidently distinguished, and the small-spored species can form five clusters that fundamentally paralleled the morphological groupings.     

Metabolic profiling of strawberry (Fragaria x ananassa Duch.) during fruit development and maturation

Strawberry (Fragaria × ananassa Duch), a fruit of economic and nutritional importance, is also a model species for fleshy fruits and genomics in Rosaceae. Strawberry fruit quality at different harvest stages is a function of the fruit's metabolite content, which results from physiological changes during fruit growth and ripening. In order to investigate strawberry fruit development, untargeted (GC-MS) and targeted (HPLC) metabolic profiling analyses were conducted. Principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA) were employed to explore the non-polar and polar metabolite profiles from fruit samples at seven developmental stages. Different cluster patterns and a broad range of metabolites that exerted influence on cluster formation of metabolite profiles were observed. Significant changes in metabolite levels were found in both fruits turning red and fruits over-ripening in comparison with red-ripening fruits. The levels of free amino acids decreased gradually before the red-ripening stage, but increased significantly in the over-ripening stage. Metabolite correlation and network analysis revealed the interdependencies of individual metabolites and metabolic pathways. Activities of several metabolic pathways, including ester biosynthesis, the tricarboxylic acid cycle, the shikimate pathway, and amino acid metabolism, shifted during fruit growth and ripening. These results not only confirmed published metabolic data but also revealed new insights into strawberry fruit composition and metabolite changes, thus demonstrating the value of metabolomics as a functional genomics tool in characterizing the mechanism of fruit quality formation, a key developmental stage in most economically important fruit crops.