A new method for the detection of carotenoids in chlorophyll samples

Summary A new method is developed for the detection of carotenoids in chlorophyll samples.The typical colour curve of chlorophyll in 80 % methyl alcohol exhibits marked absorption in 690 to 610 and less absorption in 500 to 430. The colour curve of chlorophyll contaminated with carotenoids exhibits higher values in the region 530 to 430. If carotin is present, the band maximum in the region 530 to 430 is located at 500, and if xanthophyll is the impurity the band maximum is shifted to 430. On a comparison of the colour curve of the sample to be tested with that of the typical colour curve of chlorophyll the presence of carotenoids at once becomes evident, and carotin and xanthophyll are identified separately by the positions of the band maxima in the region 500 to 430.Carotenoids in as low a concentration as 0.05 % are detected by the new method described.     

A new method for mite extraction from leaf samples

Debris often hampers the detection of mites in washed leaf samples. We describe in detail a method for the extraction of mites from leaf samples, based on the adherence of mite cuticles to liquid paraffin, at the interface of paraffin and ethanol in a so-called mite-counting channel. We demonstrate its efficacy by comparing the mite numbers in samples before and after extraction. We illustrate the method's reliability by extracting known numbers of a taxonomic variety of plant-inhabiting mites, manually added to mite-free debris: for 13 of the 15 taxa all mites were retrieved. This method can also be used to extract small non-mite arthropods such as scales, whiteflies, thrips, cicadellids, hymenopteran parasitoids and psyllids.     

基于因子分析的苜蓿叶片叶绿素高光谱反演研究

因子分析是一种能够将具有错综复杂关系的变量归结为少数几个综合因子的多变量统计分析方法,在降低数据维数的同时又可以保存足够的信息,这为处理信息量丰富但冗余较大的高光谱数据提供了一种有效方法。本文利用2010年9月23日采集的16个样点的苜蓿叶片反射率及叶绿素含量数据,采用因子分析方法,分别提取苜蓿叶片反射率光谱400～900nm,以及可见光400nm～760nm和近红外760～900nm光谱区的公共因子,分析因子载荷分布、载荷总量对公共因子与叶绿素含量相关性的影响。利用逐步回归法建立基于公共因子的叶片叶绿素反演模型,并将反演模型与光谱指数建立的模型进行对比。研究表明,1）公共因子与叶片叶绿素的相关性,在更大程度上是与该因子在各个波段上载荷分布有关,而不是总载荷量;2）对波谱进行分区建立的反演模型略优于全区因子分析建立的反演模型;3）与常用于叶片叶绿素含量反演的光谱指数CARI、MCARI、mND680、mND705、mSR705、TVI、DmSR、BGI、BRI相比,因子分析建立的叶绿素反演模型精度更高。     

A non-destructive determination of leaf chlorophyll in Vitis vinifera

A portable leaf greenness meter (SPAD-501) has been used to provide a rapid and non-destructive measurement of leaf chlorophyll in Vitis vinifera. Leaf extracted chlorophyll was related linearly to SPAD readings. It is suggested that separate linear equations should be developed for each cultivar so as to maximise the accuracy of estimating leaf chlorophyll content as a function of SPAD readings.     

Interlaboratory variability in the chemical analysis of leaf samples

Summary Statistical analyses of the results of an inter-laboratory trial on the chemical analysis of the same leaf materials have revealed significant differences between the laboratories and within the same laboratories in the case of duplicate samples analysed at the same or different times. Ranking of laboratories according to the deviation of the results from mean values suggests real differences in the consistency of the methods used.The first author was head of the Statistical Section of the Forest Research Station De Dorschkamp at Wageningen, Holland at the time of his research; the second author is senior silviculturist at the same institute and leader of the working party S1.02–3 of IUFRO (International Union of Forest Research Organizations).     

MODIS植被指数与水稻叶面积指数及叶片叶绿素含量相关性研究

利用光谱分辨率为3nm的ASD FieldSpec UV／VNIR光谱仪获得了2002和2003年水稻整个生长期的高光谱数据,同时对水稻叶面积指数(LAI)和叶绿素含量(CHL．Ｃ)进行了测定,对中分辨率成像光谱仪(MODIS)的增强植被指数(EVI)、归一化植被指数(NＤVI)以及红边位置(REP)与LAI及CHL．C之间的关系进行了研究．结果表明。LAI与冠层光谱在可见光、近红外波段相关性较好,叶绿素含量与冠层光谱在红光波段相关性较好．EVI、REP和LAI之间的相关关系不受水稻覆盖率的影响;NDVI与LAI的相关关系在水稻低覆盖率情况下较好;在水稻高覆盖率情况下。EVI和REP比NDVI与LAI之间的相关关系要好．MODIS-NDVI、EVI及REP与叶片叶绿素含量相关性较好．由此可见,EVI和REP可以有效地监测水稻LAI和CHL．C．     

Ice-cap. A high-throughput method for capturing plant tissue samples for genotype analysis

High-throughput genotype screening is rapidly becoming a standard research tool in the post-genomic era. A major bottleneck currently exists, however, that limits the utility of this approach in the plant sciences. The rate-limiting step in current high-throughput pipelines is that tissue samples from living plants must be collected manually, one plant at a time. In this article I describe a novel method for harvesting tissue samples from living seedlings that eliminates this bottleneck. The method has been named Ice-Cap to reflect the fact that ice is used to capture the tissue samples. The planting of seeds, growth of seedlings, and harvesting of tissue are all performed in a 96-well format. I demonstrate the utility of this system by using tissue harvested by Ice-Cap to genotype a population of Arabidopsis seedlings that is segregating a previously characterized mutation. Because the harvesting of tissue is performed in a nondestructive manner, plants with the desired genotype can be transferred to soil and grown to maturity. I also show that Ice-Cap can be used to analyze genomic DNA from rice (Oryza sativa) seedlings. It is expected that this method will be applicable to high-throughput screening with many different plant species, making it a useful technology for performing marker assisted selection.     

A method for quantitative analysis of spatially variable physiological processes across leaf surfaces

Many physiological processes are spatially variable across leaf surfaces. While maps of photosynthesis, stomatal conductance, gene expression, water transport, and the production of reactive oxygen species (ROS) for individual leaves are readily obtained, analytical methods for quantifying spatial heterogeneity and combining information gathered from the same leaf but with different instruments are not widely used. We present a novel application of tools from the field of geographical imaging to the multivariate analysis of physiological images. Procedures for registration and resampling, cluster analysis, and classification provide a general framework for the analysis of spatially resolved physiological data. Two experiments were conducted to illustrate the utility of this approach. Quantitative analysis of images of chlorophyll fluorescence and the production of ROS following simultaneous exposure of soybean leaves to atmospheric O₃ and soybean mosaic virus revealed that areas of the leaf where the operating quantum efficiency of PSII was depressed also experienced an accumulation of ROS. This correlation suggests a causal relationship between oxidative stress and inhibition of photosynthesis. Overlaying maps of leaf surface temperature and chlorophyll fluorescence following a photoinhibition treatment indicated that areas with low operating quantum efficiency of PSII also experienced reduced stomatal conductance (high temperature). While each of these experiments explored the covariance of two processes by overlaying independent images gathered with different instruments, the same procedures can be used to analyze the covariance of information from multiple images. The application of tools from geographic image analysis to physiological processes occurring over small spatial scales will help reveal the mechanisms generating spatial variation across leaves.     

A novel method for the transport and long-term storage of cultures and samples in an anaerobic atmosphere

Nylon-aluminium-polyethylene laminated, sealable bags provided a more economical and flexible method of transportation, incubation and storage than anaerobic jars. The bags were used with a special heat sealer and 'Anaerocult-A' sachets to remove oxygen. The environment inside the bags remained oxygen-free for at least 3 years.     

A novel method for the estimation of soybean chlorophyll content using a smartphone and image analysis

The development of smartphones, specifically their cameras, and imaging technologies has enabled their use as sensors/measurement tools. Here we aimed to evaluate the applicability of a fast and noninvasive method for the estimation of total chlorophyll (Chl), Chl a, Chl b, and carotenoids (Car) content of soybean plants using a smartphone camera. Single leaf disc images were obtained using a smartphone camera. Subsequently, for the same leaf discs, a Chl meter was used to obtain the relative index of Chl and the photosynthetic pigments were then determined using a classic method. The RGB, HSB and CIELab color models were extracted from the smartphone images and correlated to Chl values obtained using a Chl meter and by a standard laboratory protocol. The smartphone camera was sensitive enough to capture successfully a broad range of Chl and Car contents seen in soybean leaves. Although there was a variation between color models, some of the proposed regressions (e.g., the S and b index from HSB and Lab color models and NRI [RGB model]) were very close to the Chl meter values. Based on our findings, smartphones can be used for rapid and accurate estimation of soybean and Car contents in soybean leaves.     

A method for wet-ashing small plant samples

Summary An inexpensive but efficient and fast method is described for the wetashing of small samples of plant materials.Contribution No. 582, Department of Botany and Plant Pathology, Kansas Agricultural Experiment Station. Botany serial No. 744.     

Leaf chlorophyll content as a proxy for leaf photosynthetic capacity

Improving the accuracy of estimates of forest carbon exchange is a central priority for understanding ecosystem response to increased atmospheric CO₂ levels and improving carbon cycle modelling. However, the spatially continuous parameterization of photosynthetic capacity (Vcmax) at global scales and appropriate temporal intervals within terrestrial biosphere models (TBMs) remains unresolved. This research investigates the use of biochemical parameters for modelling leaf photosynthetic capacity within a deciduous forest. Particular attention is given to the impacts of seasonality on both leaf biophysical variables and physiological processes, and their interdependent relationships. Four deciduous tree species were sampled across three growing seasons (2013–2015), approximately every 10 days for leaf chlorophyll content (Chl_Leaf) and canopy structure. Leaf nitrogen (N_Area) was also measured during 2014. Leaf photosynthesis was measured during 2014–2015 using a Li‐6400 gas‐exchange system, with A‐Ci curves to model Vcmax. Results showed that seasonality and variations between species resulted in weak relationships between Vcmax normalized to 25°C () and N_Area (R² = 0.62, P < 0.001), whereas Chl_Leaf demonstrated a much stronger correlation with (R² = 0.78, P < 0.001). The relationship between Chl_Leaf and N_Area was also weak (R² = 0.47, P < 0.001), possibly due to the dynamic partitioning of nitrogen, between and within photosynthetic and nonphotosynthetic fractions. The spatial and temporal variability of was mapped using Landsat TM/ETM satellite data across the forest site, using physical models to derive Chl_Leaf. TBMs largely treat photosynthetic parameters as either fixed constants or varying according to leaf nitrogen content. This research challenges assumptions that simple N_Area– relationships can reliably be used to constrain photosynthetic capacity in TBMs, even within the same plant functional type. It is suggested that Chl_Leaf provides a more accurate, direct proxy for and is also more easily retrievable from satellite data. These results have important implications for carbon modelling within deciduous ecosystems.     

A method for selective enrichment and analysis of nitrotyrosine-containing peptides in complex proteome samples

Elevated levels of protein tyrosine nitration have been found in various neurodegenerative diseases and age-related pathologies. Until recently, however, the lack of an efficient enrichment method has prevented the analysis of this important low-level protein modification. We have developed a method that specifically enriches nitrotyrosine-containing peptides so that both nitrotyrosine peptides and specific nitration sites can be unambiguously identified with LC-MS/MS. The procedure consists of the derivatization of nitrotyrosine into free sulfhydryl groups followed by high efficiency enrichment of sulfhydryl-containing peptides with thiopropyl sepharose beads. The derivatization process includes: (1) acetylation with acetic anhydride to block all primary amines, (2) reduction of nitrotyrosine to aminotyrosine, (3) derivatization of aminotyrosine with N-Succinimidyl S-Acetylthioacetate (SATA), and (4) deprotection of S-acetyl on SATA to form free sulfhydryl groups. The high specificity of this method is demonstrated by the contrasting percentage of nitrotyrosine-derivatized peptides in the identified tandem mass spectra between enriched and unenriched samples. Global analysis of unenriched in vitro nitrated human histone H1.2, bovine serum albumin (BSA), and mouse brain homogenate samples had 9%, 9%, and 5.9% of identified nitrotyrosine-containing peptides, while the enriched samples had 91% , 62%, and 35%, respectively. Duplicate LC-MS/MS analyses of the enriched mouse brain homogenate identified 150 unique nitrated peptides covering 102 proteins with an estimated 3.3% false discovery rate.     

A sensitive and convenient method for lipoprotein profile analysis of individual mouse plasma samples

A simple and convenient method to determine plasma cholesterol profiles in individual mouse plasma samples is not presently available. With commonly used methods, plasma samples from several animals in a study group must often be pooled and analyzed, usually by the fast phase liquid chromatography (FPLC) method. The Column Lipoprotein Profile (or CLiP) method described here is a modification of the FPLC method that provides a simple and convenient procedure for determining plasma lipoprotein cholesterol profiles in small sample volumes, allowing determination of profiles from individual animals rather than from pooled plasma. The CLiP method is reproducible; a human sample measured five times over several days produced coefficients of variation as follows: VLDL, 10.0%; LDL, 0.93%; and HDL, 2.51%. CLiP-derived total cholesterol values of five different human samples (with total cholesterol levels ranging from 198 to 263 mg/dL) differed from VAP-II by -1.88% +/- 2.57%. Linearity of differing concentrations for each of the lipoprotein classes was determined by measuring the same sample with different aliquot sizes. The linear regression from VLDL had an r value of 0.996, while LDL, HDL, and total cholesterol all had r values of greater than 0.999. We present a direct comparison of plasma cholesterol profiles from several mouse models with gene modification or expression of transgenic proteins.In conclusion, the CLiP method provides a simple, reliable, and reproducible procedure for determination of plasma cholesterol profiles from individual plasma samples with very low sample volumes, using readily available equipment and reagents.     

A new optical leaf-clip meter for simultaneous non-destructive assessment of leaf chlorophyll and epidermal flavonoids

We have characterized a new commercial chlorophyll (Chl) and flavonoid (Flav) meter called Dualex 4 Scientific (Dx4). We compared this device to two other Chl meters, the SPAD-502 and the CCM-200. In addition, Dx4 was compared to the leaf-clip Dualex 3 that measures only epidermal Flav. Dx4 is factory-calibrated to provide a linear response to increasing leaf Chl content in units of μg cm(-2) , as opposed to both SPAD-502 and CCM-200 that have a non-linear response to leaf Chl content. Our comparative calibration by Chl extraction confirmed these responses. It seems that the linear response of Dx4 derives from the use of 710 nm as the sampling wavelength for transmittance. The major advantage of Dx4 is its simultaneous assessment of Chl and Flav on the same leaf spot. This allows the generation of the nitrogen balance index (NBI) used for crop surveys and nitrogen nutrition management. The Dx4 leaf clip, that incorporates a GPS receiver, can be useful for non-destructive estimation of leaf Chl and Flav contents for ecophysiological research and ground truthing of remote sensing of vegetation. In this work, we also propose a consensus equation for the transformation of SPAD units into leaf Chl content, for general use.     

A leaf disc method for measuring cuticular conductance

This paper describes a new method for the measurement of cuticularconductance (g_o;) using a leaf disc sealed in a specially-designedenvelope. Conductances for astomatous (adaxial) and stomatous(abaxial) surfaces of beech {Fagus sylvatica L.) were determinedfrom measurements of water flux. Leaf discs were punched outfrom attached leaves and placed inside individual envelopesthat provided a water supply. Water flux from an exposed epidermalsurface of the leaf discs was measured gravimetrically. Allmeasurements were made under darkness. Conductance of the adaxialsurface was referred to as g_c, whereas conductance of the abaxialsurface was considered as a minimum leaf surface conductance()- The main advantage of this method is that it enables measurement of g_c and from leaf samples with intact cuticles and a highrelative water content [RWC) for periods of up to 12 d. Conductancesof leaf discs in envelopes were compared with those of wholeleaves and leaf discs without envelopes. Data demonstratinga strong positive relationship between conductance and RWC ispresented. Key words: Cuticular conductance, leaf disc, relative water content