Cloning of genes differentially expressed during the initial stage of fruit development in melon (Cucumis melo cv. Reticulatus)

We have cloned genes involved in the initial stage of fruit development in the melon by suppression subtractive hybridization. A cDNA library of unfertilized ovules was subtracted from that of fruit 9 days after pollination (DAP); 10 of the 40 selected cDNA clones were identified by reverse Northern analysis as genes differentially expressed in fruit at 9 DAP. Seven of the ten genes were homologous to genes of known function; two were related to genes with unknown functions, and one was novel. With the exception of cucumisin, none of the cDNAs had been previously identified in melon. According to Northern analyses, six of the genes were expressed at high levels early in fruit development. Expression of cucumisin, Cmf-25, Cmf-30, and Cmf-124 was highest at 9 DAP, implying that these genes are involved in the initial stage of fruit development. Cmf-30, a seed nucellus-specific gene, was also expressed early in seed development. The other genes were expressed at a moderate level throughout fruit development, with the highest expression occurring in fruit at 9 and 18 DAP. In conclusion, nine new genes involved in early fruit development in melon were cloned, and their temporal and spatial expression patterns indicate that they are preferentially expressed during the active growing stage of fruit.     

Carbohydrate availability modifies sorbitol dehydrogenase activity of apple fruit

Complete defoliation of the stem or spur and girdling the phloem (D/G) subtending a single apple ( Malus domestica Borkh.) fruit at 70 or more days after bloom resulted in reduction of fruit growth. Extractable sorbitol dehydrogenase (SDH) (enzyme code, 1.1.1.14) activity and sorbitol and starch content of the cortex tissue of fruit receiving D/G treatment subsequently declined. Co-extraction of control and D/G cortex tissues yielded expected levels of SDH activity, indicating that the loss of extractable SDH activity in D/G fruit was not due to presence of an inhibitor or proteolytic activity. Incubating cortex sections from D/G fruit in a buffered 200 m M sorbitol or glucose solution increased extractable SDH activity, and incubating cortex sections from control fruit in the sorbitol solution maintained the activity. However, neither 200 m M fructose or 27 m M PEG, the latter with the same osmotic potential as the sorbitol solution, affected extractable SDH activity of D/G fruit. The results indicate that carbohydrate availability may affect extractable SDH activity of apple fruit, and that specific carbohydrates such as sorbitol and glucose may be signals for modulating this activity.     

Cloning and molecular characterization of a strawberry fruit ripening-related cDNA corresponding a mRNA for a low-molecular-weight heat-shock protein

We have isolated and characterized a cDNA from a strawberry fruit subtractive library that shows homology to class-I low-molecular-weight (LMW) heat-shock protein genes from other higher plants. The strawberry cDNA (clone njjs4) was a 779 bp full-length cDNA with a single open reading frame of 468 bp that is expected to encode a protein of ca. 17.4 kDa with a pI of 6.57. Southern analysis with genomic DNA showed several high-molecular-weight hybridization bands, indicating that the corresponding njjs4 gene is not present as a single copy in the genome. This strawberry gene was not expressed in roots, leaves, flowers and stolons but in fruits at specific stages of elongation and ripening. However, a differential pattern of mRNA expression was detected in the fruit tissues achenes and receptacle. The njjs4 gene expression increased in achenes accompanying the process of seed maturation whereas in the receptacle, a high mRNA expression was detected in the W2 stage, during which most of the metabolic changes leading to the fruit ripening are occurring. Our results clearly show a specific relationship of this njjs4 strawberry gene with the processes of seed maturation and fruit ripening, and strongly support that at least some of the class-I LMW heat-shock protein-like genes have a heat-stress-independent role in plant development, including fruit ripening.     

苹果中磷酸蔗糖合酶家族基因的表达特性及其与蔗糖含量的关系

磷酸蔗糖合酶(sucrose phosphate synthase,SPS)是植物中蔗糖合成的主要限速酶,影响植物的生长发育和果实中蔗糖的含量。为探明苹果中SPS基因家族特性及其在蔗糖合成中的作用,该研究从苹果基因组中分离了MdSPS家族基因,分析了它们的进化关系以及mRNA表达特性与酶活性和蔗糖含量的关系。结果显示:(1)在苹果基因组中有8个SPS家族基因表达,它们分别属于双子叶植物的3个SPS亚家族。(2)荧光定量PCR分析显示,苹果C类的MdSPS6基因和A类的MdSPS1a/b基因是苹果中表达丰度最高的SPS基因成员,其中MdSPS6在苹果成熟果中表达丰度最高,其次是成熟叶片,而MdSPS1a/b在不积累蔗糖的幼果中表达丰度最高。(3)在果实发育过程中,除MdSPS1a/b之外,其它5个苹果MdSPS家族基因均随果实的生长表达丰度增加,与SPS活性和蔗糖含量明显呈正相关关系。研究表明,C类家族MdSPS6是苹果果实发育后期和叶片中蔗糖合成的主要SPS基因。     

Microarray analysis of differentially expressed genes engaged in fruit development between table and wine grape

Microarray analysis of genes can provide individual gene-expression profiles and new insights for elucidating biological mechanisms responsible for fruit development. To obtain an overall view on expression profiles of metabolism-related genes involved in fruit development of table and wine grapes, a microarray system comprising 15,403 ESTs was used to compare the expressed genes. The expression patterns from the microarray analysis were validated with quantitative real-time polymerase chain reaction analysis of 18 selected genes of interest. During the entire fruit development stage, 2,493 genes exhibited at least 2.0-fold differences in expression levels with 1,244 genes being up-regulated and 1,249 being down-regulated. Following gene ontology analysis, only 929 differentially expressed (including 403 up-regulated and 526 down-regulated) genes were annotated in table and wine grapes. These differentially expressed genes were found to be mainly involved in carbohydrate metabolism, biosynthesis of secondary metabolites as well as energy, lipid and amino acid metabolism via KEGG. Our results provide new insights into the molecular mechanisms and expression profiles of genes in the fruit development stage of table and wine grapes.     

胃癌差异表达基因在染色体上的定位及其功能分析

利用标准化的Affymetrix公司生产的U133A基因芯片检测胃癌（T）与切缘正常胃黏膜（C）基因表达谱差异,并利用生物信息学方法对检测结果进行差异基因在染色体定位和功能分析。结果表明：胃癌与正常胃黏膜比较差异8倍以上共有270个基因,其中表达上调[信号比的对数值（SLR）≥3]有157个,表达下调（SLR≤-3）有113个。从表达差异的基因在染色体定位分析,发现除4个基因未知其定位外,其余所有差异表达基因散在分布和各条染色体上,但以1号染色体为最多,有26个（占9．8％）,其次是11和19号染色体上分别有24个（各占9．1％）。而差异表达的基因发生在染色体短臂（q）上有173个（占65％）。从表达差异的基因功能分类看,属于酶和酶调控子基因最多（67个,24．8占％）,其次是信号传导基因（43个,占15．9％）,第3类是核酸结合基因（17个,占6．3％）,第4类是转运子基因（15个,占5．5％）,第5类是蛋白结合基因（12个,占4．4％）,还有功能未知的基因有50个,占18．5％。以上5大类共占基因总数56．9％。胃癌差异表达基因散在分布在各条染色体上,但以1、11、19号染色体差异表达基因居多。这5大类（酶和酶调控子、信号传导、核酸结合、转运子、蛋白结合）相关基因异常是今后研究胃癌的重要基因。     

Function and Chromosome Location of Differentially Expressed Genes in Gastric Cancer

Using Affymetrix U133A oligonucleotide microarrays, screening was done for genes that were differentially expressed in gastric cancer (T) and normal gastric mucosa (C), and their chromosome location was characterized by bioinformatics. A total of 270 genes were found to have a difference in expression levels of more than eight times. Of them 157 were up-regulated (Signal Log Ratio [SLR]≥3), and 113 were down-regulated (SLR≤-3). Except for, four genes with unknown localization, a vast majority of the genes were sporadically distributed over every chromosome. However, chromosome 1 contained the most differentially expressed genes (26 genes, or 9.8%), followed by chromosomes 11 and 19 (both 24 genes, or 9.1%). These genes were also more likely to be on the short-arm of the chromosome (q), which had 173 (65%). When these genes were classified according to their functions, it was found that most (67 genes, 24.8%) belonged to the enzymes and their regulators groups. The next group was the signal transduction genes group (43 genes, 15.9%). The rest of the top three groups were nucleic acid binding genes (17, 6.3%), transporter genes (15, 5.5%), and protein binding genes (12, 4.4%). These made up 56.9% of all the differentially expressed genes. There were also 50 genes of unknown function (18.5%). Therefore it was concluded that differentially expressed genes in gastric cancer seemed to be sporadically distributed across the genome, but most were found on chromosomes 1, 11 and 19. The five groups associated genes abnormality were important genes for further study on gastric cancer.     

A genome-wide analysis of the expansin genes in Malus × Domestica

Expansins were first identified as cell wall-loosening proteins; they are involved in regulating cell expansion, fruits softening and many other physiological processes. However, our knowledge about the expansin family members and their evolutionary relationships in fruit trees, such as apple, is limited. In this study, we identified 41 members of the expansin gene family in the genome of apple (Malus × Domestica L. Borkh). Phylogenetic analysis revealed that expansin genes in apple could be divided into four subfamilies according to their gene structures and protein motifs. By phylogenetic analysis of the expansins in five plants (Arabidopsis, rice, poplar, grape and apple), the expansins were divided into 17 subgroups. Our gene duplication analysis revealed that whole-genome and chromosomal-segment duplications contributed to the expansion of Mdexpansins. The microarray and expressed sequence tag (EST) data showed that 34 Mdexpansin genes could be divided into five groups by the EST analysis; they may also play different roles during fruit development. An expression model for MdEXPA16 and MdEXPA20 showed their potential role in developing fruit. Overall, our study provides useful data and novel insights into the functions and regulatory mechanisms of the expansin genes in apple, as well as their evolution and divergence. As the first step towards genome-wide analysis of the expansin genes in apple, our results have established a solid foundation for future studies on the function of the expansin genes in fruit development.