scholarly journals Selection of a marker gene to construct a reference library for wetland plants, and the application of metabarcoding to analyze the diet of wintering herbivorous waterbirds

PeerJ ◽  
2016 ◽  
Vol 4 ◽  
pp. e2345 ◽  
Author(s):  
Yuzhan Yang ◽  
Aibin Zhan ◽  
Lei Cao ◽  
Fanjuan Meng ◽  
Wenbin Xu
Keyword(s):  
Marker Gene ◽  
Pcr Amplification ◽  
Food Composition ◽  
Wetland Plants ◽  
Marker Genes ◽  
Amplification Efficiency ◽  
Reference Library ◽  
Bean Goose ◽  
Dna Metabarcoding ◽  
Almost All

Food availability and diet selection are important factors influencing the abundance and distribution of wild waterbirds. In order to better understand changes in waterbird population, it is essential to figure out what they feed on. However, analyzing their diet could be difficult and inefficient using traditional methods such as microhistologic observation. Here, we addressed this gap of knowledge by investigating the diet of greater white-fronted gooseAnser albifronsand bean gooseAnser fabalis, which are obligate herbivores wintering in China, mostly in the Middle and Lower Yangtze River floodplain. First, we selected a suitable and high-resolution marker gene for wetland plants that these geese would consume during the wintering period. Eight candidate genes were included:rbcL,rpoC1,rpoB,matK,trnH-psbA,trnL (UAA),atpF-atpH, andpsbK-psbI. The selection was performed via analysis of representative sequences from NCBI and comparison of amplification efficiency and resolution power of plant samples collected from the wintering area. ThetrnL gene was chosen at last with c/h primers, and a local plant reference library was constructed with this gene. Then, utilizing DNA metabarcoding, we discovered 15 food items in total from the feces of these birds. Of the 15 unique dietary sequences, 10 could be identified at specie level. As for greater white-fronted goose, 73% of sequences belonged toPoaceaespp., and 26% belonged toCarexspp. In contrast, almost all sequences of bean goose belonged toCarexspp. (99%). Using the same samples, microhistology provided consistent food composition with metabarcoding results for greater white-fronted goose, while 13% ofPoaceaewas recovered for bean goose. In addition, two other taxa were discovered only through microhistologic analysis. Although most of the identified taxa matched relatively well between the two methods, DNA metabarcoding gave taxonomically more detailed information. Discrepancies were likely due to biased PCR amplification in metabarcoding, low discriminating power of current marker genes for monocots, and biases in microhistologic analysis. The diet differences between two geese species might indicate deeper ecological significance beyond the scope of this study. We concluded that DNA metabarcoding provides new perspectives for studies of herbivorous waterbird diets and inter-specific interactions, as well as new possibilities to investigate interactions between herbivores and plants. In addition, microhistologic analysis should be used together with metabarcoding methods to integrate this information.

scholarly journals A plant regeneration platform to apply new breeding techniques for improving disease resistance in grapevine rootstocks and cultivars

2019 ◽  
Vol 12 ◽  
pp. 01019 ◽  
Author(s):  
S. Sabbadini ◽  
L. Capriotti ◽  
C. Limera ◽  
O. Navacchi ◽  
G. Tempesta ◽  
...  
Keyword(s):  
Marker Gene ◽  
Marker Genes ◽  
Wine Grape ◽  
Thompson Seedless ◽  
Multiple Virus ◽  
Almost All ◽  
New Breeding Techniques ◽  
Breeding Techniques ◽  
Selection Of

Worldwide grapevine cultivation is based on the use of elite cultivars, in many cases strictly linked to local important wine brands. Most of Vitis viniferacultivars have high susceptibility to fungal and viral diseases therefore, new breeding techniques (e.g. Cisgenesis, RNAi and gene editing) offer the possibility to introduce new clones of the main cultivars with increased diseases resistance, in order to reduce environmental impact and improve quality in the intensive wine grape industry. This study is finalized to develop efficient in vitro regeneration and transformation protocols to extend the application of these technologies in wine grape cultivars and rootstocks. With this aim, in vitro regeneration protocols based on the production of meristematic bulks (Mezzetti et al., 2002) were optimized for different grapevine cultivars (Glera, Vermentino, Sangiovese, Thompson Seedless) and rootstocks (1103 Paulsen, and 110 Richter). The meristematic bulks were then used as explants for Agrobacteriummediated genetic transformation protocols, by comparing the use of NPTII and e-GFP as marker genes. Results confirmed the efficiency of meristematic bulks as the regenerating tissue to produce new modified plants in almost all the above genotypes. The highest regeneration efficiency in some genotypes allowed the selection of stable modified lines/calli with only the use of e-GFP marker gene. This protocol can be applied in the use of MYB marker gene for the production of cisgenic lines. Genotypes having the highest regeneration and transformation efficiency were also used for transformation experiments using a hairpin gene construct designed to silence the RNA-dependent RNA polymerase (RpRd) of the GFLV and GLRaV3, which would induce multiple virus resistances, and the Dicer-like protein 1 (Bc-DCL1) and Bc-DCL2 to control B. cinerea infection.

scholarly journals PEMA: a flexible Pipeline for Environmental DNA Metabarcoding Analysis of the 16S/18S ribosomal RNA, ITS, and COI marker genes

GigaScience ◽  
2020 ◽  
Vol 9 (3) ◽  
Author(s):  
Haris Zafeiropoulos ◽  
Ha Quoc Viet ◽  
Katerina Vasileiadou ◽  
Antonis Potirakis ◽  
Christos Arvanitidis ◽  
...  
Keyword(s):  
High Performance Computing ◽  
Ribosomal Rna ◽  
High Performance ◽  
Marker Gene ◽  
Environmental Dna ◽  
Marker Genes ◽  
Sequence Variant ◽  
18S Ribosomal Rna ◽  
Dna Metabarcoding ◽  
Performance Computing

Abstract Background Environmental DNA and metabarcoding allow the identification of a mixture of species and launch a new era in bio- and eco-assessment. Many steps are required to obtain taxonomically assigned matrices from raw data. For most of these, a plethora of tools are available; each tool's execution parameters need to be tailored to reflect each experiment's idiosyncrasy. Adding to this complexity, the computation capacity of high-performance computing systems is frequently required for such analyses. To address the difficulties, bioinformatic pipelines need to combine state-of-the art technologies and algorithms with an easy to get-set-use framework, allowing researchers to tune each study. Software containerization technologies ease the sharing and running of software packages across operating systems; thus, they strongly facilitate pipeline development and usage. Likewise programming languages specialized for big data pipelines incorporate features like roll-back checkpoints and on-demand partial pipeline execution. Findings PEMA is a containerized assembly of key metabarcoding analysis tools that requires low effort in setting up, running, and customizing to researchers’ needs. Based on third-party tools, PEMA performs read pre-processing, (molecular) operational taxonomic unit clustering, amplicon sequence variant inference, and taxonomy assignment for 16S and 18S ribosomal RNA, as well as ITS and COI marker gene data. Owing to its simplified parameterization and checkpoint support, PEMA allows users to explore alternative algorithms for specific steps of the pipeline without the need of a complete re-execution. PEMA was evaluated against both mock communities and previously published datasets and achieved results of comparable quality. Conclusions A high-performance computing–based approach was used to develop PEMA; however, it can be used in personal computers as well. PEMA's time-efficient performance and good results will allow it to be used for accurate environmental DNA metabarcoding analysis, thus enhancing the applicability of next-generation biodiversity assessment studies.

scholarly journals Crayfish hemocytes develop along the granular cell lineage

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Fang Li ◽  
Zaichao Zheng ◽  
Hongyu Li ◽  
Rongrong Fu ◽  
Limei Xu ◽  
...  
Keyword(s):  
Developmental Stages ◽  
Cell Lineage ◽  
Granular Cell ◽  
Marker Genes ◽  
Hematopoietic Tissue ◽  
Differentiated Cells ◽  
Stage Of Development ◽  
Almost All ◽  
Relationship Of

AbstractDespite the central role of hemocytes in crustacean immunity, the process of hemocyte differentiation and maturation remains unclear. In some decapods, it has been proposed that the two main types of hemocytes, granular cells (GCs) and semigranular cells (SGCs), differentiate along separate lineages. However, our current findings challenge this model. By tracking newly produced hemocytes and transplanted cells, we demonstrate that almost all the circulating hemocytes of crayfish belong to the GC lineage. SGCs and GCs may represent hemocytes of different developmental stages rather than two types of fully differentiated cells. Hemocyte precursors produced by progenitor cells differentiate in the hematopoietic tissue (HPT) for 3 ~ 4 days. Immature hemocytes are released from HPT in the form of SGCs and take 1 ~ 3 months to mature in the circulation. GCs represent the terminal stage of development. They can survive for as long as 2 months. The changes in the expression pattern of marker genes during GC differentiation support our conclusions. Further analysis of hemocyte phagocytosis indicates the existence of functionally different subpopulations. These findings may reshape our understanding of crustacean hematopoiesis and may lead to reconsideration of the roles and relationship of circulating hemocytes.

scholarly journals Multi-view feature selection for identifying gene markers: a diversified biological data driven approach

2020 ◽  
Vol 21 (S18) ◽  
Author(s):  
Sudipta Acharya ◽  
Laizhong Cui ◽  
Yi Pan
Keyword(s):  
Gene Expression ◽  
Feature Selection ◽  
Gene Selection ◽  
Marker Gene ◽  
Biological Data ◽  
Protein Interaction Data ◽  
Marker Genes ◽  
Data Sets ◽  
Gene Markers ◽  
Multi Objective

Abstract Background In recent years, to investigate challenging bioinformatics problems, the utilization of multiple genomic and proteomic sources has become immensely popular among researchers. One such issue is feature or gene selection and identifying relevant and non-redundant marker genes from high dimensional gene expression data sets. In that context, designing an efficient feature selection algorithm exploiting knowledge from multiple potential biological resources may be an effective way to understand the spectrum of cancer or other diseases with applications in specific epidemiology for a particular population. Results In the current article, we design the feature selection and marker gene detection as a multi-view multi-objective clustering problem. Regarding that, we propose an Unsupervised Multi-View Multi-Objective clustering-based gene selection approach called UMVMO-select. Three important resources of biological data (gene ontology, protein interaction data, protein sequence) along with gene expression values are collectively utilized to design two different views. UMVMO-select aims to reduce gene space without/minimally compromising the sample classification efficiency and determines relevant and non-redundant gene markers from three cancer gene expression benchmark data sets. Conclusion A thorough comparative analysis has been performed with five clustering and nine existing feature selection methods with respect to several internal and external validity metrics. Obtained results reveal the supremacy of the proposed method. Reported results are also validated through a proper biological significance test and heatmap plotting.

scholarly journals Corrigendum to: PEMA: a flexible pipeline for environmental DNA metabarcoding analysis of the 16S/18S ribosomal RNA, ITS, and COI marker genes

GigaScience ◽  
2020 ◽  
Vol 9 (12) ◽  
Author(s):  
Haris Zafeiropoulos ◽  
Ha Quoc Viet ◽  
Katerina Vasileiadou ◽  
Antonis Potirakis ◽  
Christos Arvanitidis ◽  
...  
Keyword(s):  
Ribosomal Rna ◽  
Environmental Dna ◽  
Marker Genes ◽  
18S Ribosomal Rna ◽  
Dna Metabarcoding

PPIT: an R package for inferring microbial taxonomy from nifH sequences

2021 ◽  
Author(s):  
Bennett J Kapili ◽  
Anne E Dekas
Keyword(s):  
Gene Transfer ◽  
Horizontal Gene Transfer ◽  
Query Sequence ◽  
Marker Gene ◽  
R Package ◽  
Supplementary Information ◽  
Marker Genes ◽  
Pairwise Identity ◽  
Metabolic Marker ◽  
Microbial Taxonomy

Abstract Motivation Linking microbial community members to their ecological functions is a central goal of environmental microbiology. When assigned taxonomy, amplicon sequences of metabolic marker genes can suggest such links, thereby offering an overview of the phylogenetic structure underpinning particular ecosystem functions. However, inferring microbial taxonomy from metabolic marker gene sequences remains a challenge, particularly for the frequently sequenced nitrogen fixation marker gene, nitrogenase reductase (nifH). Horizontal gene transfer in recent nifH evolutionary history can confound taxonomic inferences drawn from the pairwise identity methods used in existing software. Other methods for inferring taxonomy are not standardized and require manual inspection that is difficult to scale. Results We present Phylogenetic Placement for Inferring Taxonomy (PPIT), an R package that infers microbial taxonomy from nifH amplicons using both phylogenetic and sequence identity approaches. After users place query sequences on a reference nifH gene tree provided by PPIT (n = 6317 full-length nifH sequences), PPIT searches the phylogenetic neighborhood of each query sequence and attempts to infer microbial taxonomy. An inference is drawn only if references in the phylogenetic neighborhood are: (1) taxonomically consistent and (2) share sufficient pairwise identity with the query, thereby avoiding erroneous inferences due to known horizontal gene transfer events. We find that PPIT returns a higher proportion of correct taxonomic inferences than BLAST-based approaches at the cost of fewer total inferences. We demonstrate PPIT on deep-sea sediment and find that Deltaproteobacteria are the most abundant potential diazotrophs. Using this dataset we show that emending PPIT inferences based on visual inspection of query sequence placement can achieve taxonomic inferences for nearly all sequences in a query set. We additionally discuss how users can apply PPIT to the analysis of other marker genes. Availability PPIT is freely available to non-commercial users at https://github.com/bkapili/ppit. Installation includes a vignette that demonstrates package use and reproduces the nifH amplicon analysis discussed here. The raw nifH amplicon sequence data have been deposited in the GenBank, EMBL, and DDBJ databases under BioProject number PRJEB37167. Supplementary information Supplementary data are available at Bioinformatics online.

Recombination and deletion of sequences in shuttle vector plasmids in mammalian cells

1985 ◽  
Vol 5 (9) ◽  
pp. 2265-2271
Author(s):  
S Chakrabarti ◽  
S Joffe ◽  
M M Seidman
Keyword(s):  
Shuttle Vector ◽  
Marker Gene ◽  
Simian Virus 40 ◽  
Repeated Sequences ◽  
Marker Genes ◽  
Insertion Mutagenesis ◽  
Direct Repeats ◽  
African Green Monkey Kidney ◽  
Alu Sequence ◽  
Insertion Mutants

Shuttle vector plasmids were constructed with directly repeated sequences flanking a marker gene. African green monkey kidney (AGMK) cells were infected with the constructions, and after a period of replication, the progeny plasmids were recovered and introduced into bacteria. Those colonies with plasmids that had lost the marker gene were identified, and the individual plasmids were purified and characterized by restriction enzyme digestion. Recombination between the repeated elements generated a plasmid with a precise deletion and a characteristic restriction pattern, which distinguished the recombined molecules from those with other defects in the marker gene. Recombination among the following different sequences was measured in this assay: (i) the simian virus 40 origin and enhancer region, (ii) the AGMK Alu sequence, and (iii) a sequence from plasmid pBR322. Similar frequencies of recombination among these sequences were found. Recombination occurred more frequently in Cos1 cells than in CV1 cells. In these experiments, the plasmid population with defective marker genes consisted of the recombined molecules and of the spontaneous deletion-insertion mutants described earlier. The frequency of the latter class was unaffected by the presence of the option for recombination represented by the direct repeats. Both recombination and deletion-insertion mutagenesis were stimulated by double-strand cleavage between the repeated sequences and adjacent to the marker, and the frequency of the deletion-insertion mutants in this experiment was again independent of the presence of the direct repeats. We concluded that although recombination and deletion-insertion mutagenesis were both stimulated by double-strand cleavage, the molecules which underwent the two types of change were drawn from separate pools.

scholarly journals Method Comparison of DNA Isolation and Quantification for Fish and Seafood Authenticity Determination

2018 ◽  
Vol 13 (3) ◽  
pp. 115
Author(s):  
Dwiyitno Dwiyitno ◽  
Stefan Hoffman ◽  
Koen Parmentier ◽  
Chris Van Keer
Keyword(s):  
Polymerase Chain Reaction ◽  
Dna Extraction ◽  
Dna Isolation ◽  
Blue Mussel ◽  
Pcr Amplification ◽  
Amplification Efficiency ◽  
Chain Reaction ◽  
Seafood Products ◽  
Polymerase Chain ◽  
Commercial Kit

Fish and seafood products has been commonly targeted for fraudulent activities. For that reason, authentication of fish and seafood products is important to protect consumers from fraudulent and adulteration practices, as well as to implement traceability regulation. From the viewpoint of food safety, authenticity is beneficial to protect public from serious food poisoning incidents, such as due to ingestion of toxic species. Since DNA based identification depends on the nucleic acid polymerase chain reaction (PCR), the quantity and quality/purity of DNA will contribute significantly to the species authentication. In the present study, different DNA extraction and purification methods (3 classical methods and one commercial kit) were compared to produce the better isolated DNA for PCR amplification. Additionally, different methods for the estimation of DNA concentration and purity which is essential for PCR amplification efficiency were also evaluated. The result showed that classical DNA extraction methods (based on TNES-Urea) yielded a higher amount of DNA (11.30-323.60 ng/g tissue) in comparison to commercial kit/Wizard Promega (5.70-83.45 ng/g tissue). Based on the purity of DNA extract (A260/280), classical DNA extraction method produced relatively similar on DNA quality to the commercial kit (1.79-2.12). Interestingly, all classical methods produced DNA with A260/280 ratio of more than 2.00 on the blue mussel, in contrast with commercial kit. The commercial kit also produced better quality of DNA compared to the classical methods, showing the higher efficiency in PCR amplification. NanoDrop is promising as cheap, robust and safe UV-spectrophotometer method for DNA quantification, as well as the purity evaluation.Keywords: seafood authenticity, DNA isolation, polymerase chain reaction, NanoDrop, Picogreen

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