Barcode High Resolution Melting (Bar-HRM) analysis for detection and quantification of PDO “Fava Santorinis” (Lathyrus clymenum) adulterants

Legumes considered as one of the most important crops worldwide. Due to high price as a PDO product, commercial products of “Fava Santorinis” are often subjected to adulterations from other legume products coming from other Lathyrus or Vicia and Pisum species. Using plant DNA barcoding regions (trnL and rpoC) coupled with High Resolution Melting (Bar-HRM) we have developed a method allowing us to detect and authenticate PDO “Fava Santorinis”. Bar-HRM proved to be a very sensitive tool able to genotype Lathyrus and its closed relative species and to detect admixtures, being sensitive enough to as low as 1:100 of non-“Fava Santorinis” in “Fava Santorinis” commercial products. In conclusion, Bar-HRM analysis can be a faster, with higher resolution and cost effectiveness alternative method to authenticate PDO “Fava Santorinis” and to quantitatively detect adulterations in “Fava Santorinis” with other relative commercial “Fava” food products.     

Introducing pyruvate oxidase into the chloroplast of Chlamydomonas reinhardtii increases oxygen consumption and promotes hydrogen production

In an anaerobic environment, the unicellular green algae Chlamydomonas reinhardtii can produce hydrogen (H₂) using hydrogenase. The activity of hydrogenase is inhibited at the presence of molecular oxygen, forming a major barrier for large scale production of hydrogen in autotrophic organisms. In this study, we engineered a novel pathway to consume oxygen and correspondingly promote hydrogen production in Chlamydomonas reinhardtii. The pyruvate oxidase from Escherichia coli and catalase from Synechococcus elongatus PCC 7942 were cloned and integrated into the chloroplast of Chlamydomonas reinhardtii. These two foreign genes are driven by a HSP70A/RBCS2 promoter, a heat shock inducing promoter. After continuous heat shock treatments, the foreign genes showed high expression levels, while the growth rate of transgenic algal cells was slightly inhibited compared to the wild type. Under low light, transgenic algal cells consumed more oxygen than wild type. This resulted in lower oxygen content in sealed culture conditions, especially under low light condition, and dramatically increased hydrogen production. These results demonstrate that pyruvate oxidase expressed in Chlamydomonas reinhardtii increases oxygen consumption and has potential for improving photosynthetic hydrogen production in Chlamydomonas reinhardtii.     

A rapid screening for adulterants in olive oil using DNA barcodes

A distinctive methodology is developed to trace out the mixing into olive oil, which is marketed every year with 20% or more fraudulent oils. Such adulteration has been difficult to differentiate using fatty acid analysis and other available current techniques, as chemically fatty acids are same regardless of their source. The total genomic DNA isolated from olive oil, contaminated with canola and sunflower was analysed for single nucleotide polymorphism (SNP) variation in noncoding spacer region between psbA-trnH and partial coding region of matK of plastid genome. These DNA regions were amplified by PCR using specific primers and resulting DNA sequences were matched to the predetermined consensus DNA barcode sequences of canola and sunflower for discerning the contaminations in olive oil samples. The matching of an adulterant DNA sequence with their respective DNA barcode revealed the mixing of canola and sunflower oil into olive is simpler way and the combined approach of molecular biology and bioinformatics technology can be used as an inexpensive method for ensuring the purity of olive. This plastid based molecular DNA technology can be used for rapid detection of adulteration easily up to 5% in olive oil.     

晾制期间白肋烟TN90叶绿体色素降解动态及呼吸强度的变化

通过田间试验,研究了晾制期间白肋烟TN90叶绿体色素的降解动态及呼吸强度、含水率、比叶重等的变化。结果表明,晾制期间TN90叶绿素(chl)、叶绿素a(chla)、叶绿素b(chlb)、类胡萝卜素等的含量均随叶位的升高而增加、随晾制进程的推进而下降,其降幅chla>chl>chlb>类胡萝卜素,下、中、上各叶位chla降幅最大的时段分别为0～6d、0～9d、0～12d,其余各类色素的降低幅度则比较平缓;chla/chlb及chl/类胡萝卜素的比值也随晾制进程的推进而下降,这有利于烟叶品质的改善。随着晾制过程中水分的散失,烟叶含水率逐渐下降,叶细胞生命活动也随之减弱,表现为呼吸强度和比叶重也随晾制进程及叶位下降而减小。     

油菜叶绿体定点转化载体的构建及其杀虫性

首先从油菜叶绿体基因组克隆得到包含ｒｐｓ７基因在内的１．０ｋｂＤＮＡ片 包含ｎｄｈＢ基因在内的２．４ｋｂＤＮＡ片段,同时从苏云芽孢杆菌质粒上克隆得到一个全长３．５ｋｂ的ＢＴ杀虫蛋白基因ｃｒｙ１Ａａ。然后以ｒｐｓ７和ｎｄｈＢ基因作为同源重组片段,成功构建了包含Ｂｔ杀虫蛋白和ａａｄＡ抗壮观霉素基因的油菜叶绿体定点转化载体,并对克隆菌体总蛋白进行了生物杀虫试验。结果表明,Ｂｔ杀虫蛋白基因能够得到表达,并     

Insights into cadmium induced physiological and ultra-structural disorders in Juncus effusus L. and its remediation through exogenous citric acid

This study appraised cadmium (Cd) toxicity stress in wetland plant Juncus effusus, and explored its potential for Cd phytoextraction through chelators (citric acid and EDTA). Cadmium altered morphological and physiological attributes of J. effusus as reflected by growth retardation. Citric acid in the presence of 100 μM Cd significantly countered Cd toxicity by improving plant growth. Elevated Cd concentrations reduced translocation factor that was increased under application of both chelators. Citric acid enhanced Cd accumulation, while EDTA reduced its uptake. Cadmium induced oxidative stress modified the antioxidative enzyme activity. Both levels of citric acid (2.5 and 5.0 mM) and lower EDTA concentration (2.5 mM) helped plants to overcome oxidative stress by enhancing their antioxidative enzyme activities. Cadmium damaged the root cells through cytoplasmic shrinkage and metal deposition. Citric acid restored structure and shape of root cells and eliminated plasmolysis; whereas, EDTA exhibited no positive effect on it. Shoot cells remained unaffected under Cd treatment alone or with citric acid except for chloroplast swelling. Only EDTA promoted starch accumulation in chloroplast reflecting its negative impact on cellular structure. It concludes that Cd and EDTA induce structural and morphological damage in J. effusus; while, citric acid ameliorates Cd toxicity stress.     

古白皮松衰老机理的研究

本文主要研究古白皮松的衰老衰弱与叶绿体的超微结构及叶绿素含量的关系，通过透射电子显微镜的观察和叶绿素含量的测定发现；古白皮树龄的差异对叶绿体结构及叶绿素含量影响不大，而植物衰老衰弱时叶绿体的变化十分明显，壮树的叶绿体结构完整，弱树的叶绿体内含巨大淀粉粒，特弱树的叶绿是多被破坏，叶绿素的含量也随树木的衰弱而降低，不同季节叶绿素的含量亦有变化。     

Citric acid enhances the phytoextraction of manganese and plant growth by alleviating the ultrastructural damages in Juncus effusus L.

Chelate-assisted phytoextraction by high biomass producing plant species enhances the removal of heavy metals from polluted environments. In this regard, Juncus effusus a wetland plant has great potential. This study evaluated the effects of elevated levels of manganese (Mn) on the vegetative growth, Mn uptake and antioxidant enzymes in J. effusus. We also studied the role of citric acid and EDTA on improving metal accumulation, plant growth and Mn toxicity stress alleviation. Three-week-old plantlets of J. effusus were subjected to various treatments in the hydroponics as: Mn (50, 100 and 500 μM) alone, Mn (500 μM) + citric acid (5 mM), and Mn (500 μM) + EDTA (5 mM). After 2 weeks of treatment, higher Mn concentrations significantly reduced the plant biomass and height. Both citric acid and EDTA restored the plant height as it was reduced at the highest Mn level. Only the citric acid (but not EDTA) was able to recover the plant biomass weight, which was also obvious from the microscopic visualization of mesophyll cells. There was a concentration dependent increase in Mn uptake in J. effusus plants, and relatively more deposition in roots compared to aerial parts. Although both EDTA and citric acid caused significant increase in Mn accumulation; however, the Mn translocation was enhanced markedly by EDTA. Elevated levels of Mn augmented the oxidative stress, which was evident from changes in the activities of antioxidative enzymes in plant shoots. Raised levels of lipid peroxidation and variable changes in the activities of antioxidant enzymes were recorded under Mn stress. Electron microscopic images revealed several modifications in the plants at cellular and sub-cellular level due to the oxidative damage induced by Mn. Changes in cell shape and size, chloroplast swelling, increased number of plastoglobuli and disruption of thylakoid were noticed. However, these plants showed a high degree of tolerance against Mn toxicity stress, and it removed substantial amounts of Mn from the media. The EDTA best enhanced the Mn uptake and translocation, while citric acid best recovered the plant growth.     

Effects of 1-MCP on chlorophyll degradation pathway-associated genes expression and chloroplast ultrastructure during the peel yellowing of Chinese pear fruits in storage

The peel yellowing is an important pigment physiological process of green fruit ripening, which mainly results from chlorophyll degradation in the fruit peel. In this work, two typical cultivars with different ripening speed, a slow ripening pear 'Emerald' (Pyrus bretschneideri Rehd. cv. Emerald) and a fast ripening 'Jingbai' (Pyrus ussuriensis Maxim. cv. Jingbai) were used to investigate the molecular mechanism of chlorophyll degradation in pear yellowing/ripening during postharvest storage. The fruits after harvest were treated with 1-methylcyclopropene (1-MCP), an ethylene action inhibitor at 1.0 μLl(-1) to determine its effect on chloroplast ultrastructure and the expression of chlorophyll degradation associated genes in peel tissues. Our results show that the pears treated with 1-MCP had a lower ethylene production rate and higher chlorophyll content compared to those of untreated fruit. The more intact chloroplasts with well-organised grana thylakoids and small plastoglobuli were maintained in the peel of 1-MCP treated fruit for up to 30 and 15 d in 'Emerald' and 'Jingbai', respectively. The expression of chlorophyll degradation associated genes: pheophorbide a oxygenase (PAO), non-yellow colouring (NYC), NYC1-like (NOL), stay-green 1(SGR1), was suppressed, while no significant change was found in chlorophyllase 1 (CHL1) and red chlorophyll catabolite reductase (RCCR) in both cultivar fruits treated with 1-MCP. These results suggest that 1-MCP can delay chlorophyll degradation by inhibiting ethylene production and suppressing the gene expression of PAO, NYC, NOL and SGR1, which are closely associated with chlorophyll catabolic pathway.