Fingerprinting of complex mixtures with the use of high performance liquid chromatography, inductively coupled plasma atomic emission spectroscopy and chemometrics

The molecular and metal profile fingerprints were obtained from a complex substance, Atractylis chinensis DC—a traditional Chinese medicine (TCM), with the use of the high performance liquid chromatography (HPLC) and inductively coupled plasma atomic emission spectroscopy (ICP-AES) techniques. This substance was used in this work as an example of a complex biological material, which has found application as a TCM. Such TCM samples are traditionally processed by the Bran, Cut, Fried and Swill methods, and were collected from five provinces in China. The data matrices obtained from the two types of analysis produced two principal component biplots, which showed that the HPLC fingerprint data were discriminated on the basis of the methods for processing the raw TCM, while the metal analysis grouped according to the geographical origin. When the two data matrices were combined into a one two-way matrix, the resulting biplot showed a clear separation on the basis of the HPLC fingerprints. Importantly, within each different grouping the objects separated according to their geographical origin, and they ranked approximately in the same order in each group. This result suggested that by using such an approach, it is possible to derive improved characterisation of the complex TCM materials on the basis of the two kinds of analytical data. In addition, two supervised pattern recognition methods, K-nearest neighbors (KNNs) method, and linear discriminant analysis (LDA), were successfully applied to the individual data matrices—thus, supporting the PCA approach.

Assessment and rationalization of genetic diversity of Papua New Guinea taro (Colocasia esculenta) using SSR DNA fingerprinting

Taro (Colocasia esculenta) accessions were collected from 15 provinces of Papua New Guinea (PNG). The collection, totalling 859 accessions was collated for characterization and a core collection of 81 accessions (10%) was established on the basis of characterization data generated on 30 agro-morphological descriptors, and DNA fingerprinting using seven SSR primers. The selection of accessions was based on cluster analysis of the morphological data enabling initial selection of 20% accessions. The 20% sample was then reduced and rationalized to 10% based on molecular data generated by SSR primers. This represents the first national core collection of any species established in PNG based on molecular markers. The core has been integrated with core from other Pacific Island countries, contributing to a Pacific regional core collection, which is conserved in vitro in the South Pacific Regional Germplasm Centre at Fiji. The core collection is a valuable resource for food security of the South Pacific region and is currently being utilized by the breeding programmes of small Pacific Island countries to broaden the genetic base of the crop.

DNA amplification fingerprinting analysis of genetic variation within Fusarium oxysporum f.sp zingiberi

Genetic variation among 29 isolates of Fusarium oxysporum f.sp. zingiberi (Foz) collected from diseased ginger rhizome in production regions throughout Queensland was analysed using DNA amplification fingerprinting (DAF). Eight isolates of other Fusarium species and/or formae speciales were included for comparative analysis. Within the Foz isolates, three haplotypes were identified based on 17 polymorphic bands generated with five primers. Two groups showed very little genetic variation (98.6% similarity), whereas the third single isolate was quite distinct in terms of its molecular profile (77.2% similarity). Genetic similarity among the Fusarium solani, F. oxysporum f.sp. lycopersici and F. oxysporum f.sp. cubense races 1, 3 and 4 isolates compared well with the published literature.

Fingerprinting environmental conditions and related stress using stable isotopic composition of rice (Oryza sativa L.) grain organic matter

Sea level rise (SLR) is a primary factor responsible for inundation of low-lying coastal regions across the world, which in turn governs the agricultural productivity. In this study, rice (Oryza sativa L.) cultivated seasonally in the Kuttanad Wetland, a SLR prone region on the southwest coast of India, were analysed for oxygen, hydrogen and carbon isotopic ratios (delta O-18, delta H-2 and delta C-13) to distinguish the seasonal environmental conditions prevalent during rice cultivation. The region receives high rainfall during the wet season which promotes large supply of fresh water to the local water bodies via the rivers. In contrast, during the dry season reduced river discharge favours sea water incursion which adversely affects the rice cultivation. The water for rice cultivation is derived from regional water bodies that are characterised by seasonal salinity variation which co-varies with the delta O-18 and delta H-2 values. Rice cultivated during the wet and the dry season bears the isotopic imprints of this water. We explored the utility of a mechanistic model to quantify the contribution of two prominent factors, namely relative humidity and source water composition in governing the seasonal variation in oxygen isotopic composition of rice grain OM. delta C-13 values of rice grain OM were used to deduce the stress level by estimating the intrinsic water use efficiency (WUEi) of the crop during the two seasons. 1.3 times higher WUE, was exhibited by the same genotype during the dry season. The approach can be extended to other low lying coastal agro-ecosystems to infer the growth conditions of cultivated crops and can further be utilised for retrieving paleo-environmental information from well preserved archaeological plant remains. (c) 2015 Elsevier Ltd. All rights reserved.

PCR-DGGE Fingerprinting Analysis of Plankton Communities and Its Relationship to Lake Trophic Status

Plankton communities in eight lakes of different trophic status near Yangtze, China were characterized by using denatured gradient gel electrophoresis (DGGE). Various water quality parameters were also measured at each collection site. Following extraction of DNA from plankton communities, 16S rRNA and 18S rRNA genes were amplified with specific primers for prokaryotes and eukaryotes, respectively; DNA profiles were developed by DGGE. The plankton community of each lake had its own distinct DNA profile. The total number of bands identified at 34 sampling stations ranged from 37 to 111. Both prokaryotes and eukaryotes displayed complex fingerprints composed of a large number of bands: 16 to 59 bands were obtained with the prokaryotic primer set; 21 to 52 bands for the eukaryotic primer set. The DGGE-patterns were analyzed in relation to water quality parameters by canonical correspondence analysis (CCA). Temperature, pH, alkalinity, and the concentration of COD, TP and TN were strongly correlated with the DGGE patterns. The parameters that demonstrated a strong correlation to the DGGE fingerprints of the plankton community differed among lakes, suggesting that differences in the DGGE fingerprints were due mainly to lake trophic status. Results of the present study suggest that PCR-DGGE fingerprinting is an effective and precise method of identifying changes to plankton community composition, and therefore could be a useful ecological tool for monitoring the response of aquatic ecosystems to environmental perturbations.

Genetic diversity of plankton community as depicted by PCR-DGGE fingerprinting and its relation to morphological composition and environmental factors in lake donghu

To collect information about the genetic diversity of the plankton community and to study how plankton respond to environmental conditions, plankton samples were collected from five stations representing different trophic levels in a shallow, eutrophic lake (Lake Donghu), and investigated by PCR-DGGE fingerprinting. A total of 100 bands (61 of 16S rDNA bands and 39 of 18S rDNA bands) were detected. The DGGE bands unique to any single station accounted for 38% of the total bands, whereas common bands detected at all five stations accounted for only 11%. Using UPGMA clustering and MDS ordination of DGGE fingerprints, stations I and II were found to initially group together into one cluster, which was later joined by station V. Stations III and IV were isolated into two separate groups of one station each. Some differences in grouping relationships were found when analysis was completed on the basis of chemical characteristics and morphological composition, with zooplankton composition showing the greatest variability. However, the most similar stations (I and II) were always initially grouped into one cluster. Moreover, stations that exhibited the same or similar trophic level (stations III and IV), but different concentrations of heavy metals, were further differentiated by the DGGE method. Results of the present study indicated that PCR-DGGE fingerprinting was more sensitive than the traditional methods, as other studies suggested. Additionally, PCR-DGGE appears to be more appropriate for diversity characterization of the plankton community, as it is more canonical, systematic, and effective. Most importantly, fingerprinting results are more convenient for the comparative analyses between different studies. Therefore, the use of the described fingerprinting analysis may provide an operable and sensitive biomonitoring approach to identify critical, and potentially negative, stress within an aquatic ecosystem.

Genetic fingerprinting of plankton community provides new insights into aquatic ecology

Over the past two decades, molecular techniques have been widely used in ecological study and molecular ecology has been one of the most important branches of ecology. Meanwhile, genetic fingerprinting analyses have significantly enhanced our knowledge of the diversity and evolutionary relations of the planktonic organisms. Compared with conventional approaches in ecological study (e. g. morphological classification), genetic fingerprinting techniques are simpler and much more effective. This review provides an overview of the principles, advantages and limitations of the commonly used DNA fingerprinting techniques in plankton research. The aim of this overview is to assess where we have been, where we are now and what the future holds for solving aquatic ecological problems with molecular-level information.

Population genetic structure of Carchesium polypinum (Ciliophora : Peritrichia) in four chinese lakes inferred from ISSR fingerprinting: High diversity but low differentiation

Although the peritrichous ciliate Carchesium polypinum is common in freshwater, its population genetic structure is largely unknown. We used inter-simple sequence repeat (ISSR) fingerprinting to analyze the genetic structure of 48 different isolates of the species from four lakes in Wuhan, central China. Using eight polymorphic primers, 81 discernible DNA fragments were detected, among which 76 (93.83%) were polymorphic, indicating high genetic diversity at the isolate level. Further, Nei's gene diversity (h) and Shannon's Information index (I) between the different isolates both revealed a remarkable genetic diversity, higher than previously indicated by their morphology. At the same time, substantial gene flow was found. So the main factors responsible for the high level of diversity within populations are probably due to conjugation (sexual reproduction) and wide distribution of swarmers. Analysis of molecular variance (AMOVA) showed that there was low genetic differentiation among the four populations probably due to common ancestry and flooding events. The cluster analysis and principal component analysis (PCA) suggested that genotypes isolated from the same lake displayed a higher genetic similarity than those from different lakes. Both analyses separated C. polypinum isolates into subgroups according to the geographical locations. However, there is only a weak positive correlation between the genetic distance and geographical distance, suggesting a minor effect of geographical distance on the distribution of genetic diversity between populations of C. polypinum at the local level. In conclusion, our studies clearly demonstrated that a single morphospecies may harbor high levels of genetic diversity, and that the degree of resolution offered by morphology as a marker for measuring distribution patterns of genetically distinct entities is too low.