Construction and characterization of a de novo draft genome of garden cress (Lepidium sativum L.).

Garden cress (Lepidium sativum L.) is a Brassicaceae crop recognized as a healthy vegetable and a medicinal plant. Lepidium is one of the largest genera in Brassicaceae, yet, the genus has not been a focus of extensive genomic research. In the present work, garden cress genome was sequenced using the long read high-fidelity sequencing technology. A de novo, draft genome assembly that spans 336.5 Mb was produced, corresponding to 88.6% of the estimated genome size and representing 90% of the evolutionarily expected orthologous gene content. Protein coding gene content was structurally predicted and functionally annotated, resulting in the identification of 25,668 putative genes. A total of 599 candidate disease resistance genes were identified by predicting resistance gene domains in gene structures, and 37 genes were detected as orthologs of heavy metal associated protein coding genes. In addition, 4289 genes were assigned as "transcription factor coding." Six different machine learning algorithms were trained and tested for their performance in classifying miRNA coding genomic sequences. Logistic regression proved the best performing trained algorithm, thus utilized for pre-miRNA coding loci identification in the assembly. Repetitive DNA analysis involved the characterization of transposable element and microsatellite contents. L. sativum chloroplast genome was also assembled and functionally annotated. Data produced in the present work is expected to constitute a foundation for genomic research in garden cress and contribute to genomics-assisted crop improvement and genome evolution studies in the Brassicaceae family.

The trnL (UAA)-trnF (GAA) intergenic spacer is a robust marker of green pea (Pisum sativum L.) adulteration in economically valuable pistachio nuts (Pistacia vera L.).

BACKGROUND: Pistachio (Pistacia vera L.) is an expensive culinary nut species; it is therefore susceptible to adulteration for economic profit. Green pea (Pisum sativum L.) kernels constitute the most common material used for adulterating chopped / ground pistachio nuts and pistachio paste. Food genomics enables the species composition of a food sample to be ascertained through DNA analysis. Accordingly, a barcode DNA genotyping approach was used to standardize a test method to identify green pea adulteration in pistachio nuts. RESULTS: The trnL (UAA)-trnF (GAA) intergenic spacer in the plastid genome was the target analyte in the present study. The barcode locus displayed a significant, discriminatory size difference between pistachio and pea, with amplicon sizes of 449 and 179 bp, respectively. Polymerase chain reaction-capillary electrophoresis (PCR-CE) analysis of the intergenic spacer resulted in the successful identification of species composition in the in-house admixtures, which contained 5% to 30% of green pea. CONCLUSION: The present work describes a fast and straightforward DNA test that identifies green pea adulteration in pistachio nuts without requiring a statistical data interpretation process. The plastid trnL (UAA)-trnF (GAA) intergenic spacer length widely varies among plant taxa, so the PCR-CE protocol that operates on the intergenic spacer holds the potential to reveal adulteration with a plethora of adulterants. The PCR-CE assay described in the present work can be adopted readily by food-quality laboratories in the public sector or the food industry as an easy and reliable method to analyze pistachio authenticity. © 2020 Society of Chemical Industry.

Transcriptomic analysis of asymptomatic and symptomatic severe Turkish patients in SARS-CoV-2 infection.

Objective: Coronavirus disease 2019 (COVID-19), leading to mild infection (MI), acute respiratory distress syndrome or death in different persons. Although the basis of these variabilities has not been fully elucidated, some possible findings have been encountered. In the present study, we aimed to reveal genes with different expression profiles by next-generation sequencing of RNA isolated from blood taken from infected patients to reveal molecular causes of different response. Methods: Two healthy, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-negative control individuals (NCI), two SARS-CoV-2-positive patients who have MI, and two patients who have critical infection (CI) were included in the study. Total RNA was extracted from blood samples and sequenced. Raw RNA-Seq data were analyzed on Galaxy platform for the identification of differentially expressed genes and their pathway involvements. Results: We found that 199 and 521 genes were downregulated in whole blood of COVID-19-positive CI patients compared to NCI and MI patients, respectively. We identified 21 gene ontology pathways commonly downregulated in CI patients compared to both NCI and MI, mostly associated with innate and adaptive immune responses. Three hundred and fifty-four and 600 genes were found to be upregulated compared to NCI and MI, respectively. Upregulated six pathways included genes that function in inflammatory response and inflammatory cytokine release. Conclusion: The transcriptional profile of CI patients deviates more significantly from that of MI in terms of the number of differentially expressed genes, implying that genotypic differences may account for the severity of SARS-CoV-2 infection and inflammatory responses through differential regulation of gene expression. Therefore, further studies that involve whole genome analysis coupled with differential expression analysis are required in order to determine the dynamics of genotype - gene expression profile associations.

De novo assembly and characterization of the first draft genome of quince (Cydonia oblonga Mill.).

Quince (Cydonia oblonga Mill.) is the sole member of the genus Cydonia in the Rosacea family and closely related to the major pome fruits, apple (Malus domestica Borkh.) and pear (Pyrus communis L.). In the present work, whole genome shotgun paired-end sequencing was employed in order to assemble the first draft genome of quince. A genome assembly that spans 488.4 Mb of sequence corresponding to 71.2% of the estimated genome size (686 Mb) was produced in the study. Gene predictions via ab initio and homology-based sequence annotation strategies resulted in the identification of 25,428 and 30,684 unique putative protein coding genes, respectively. 97.4 and 95.6% of putative homologs of Arabidopsis and rice transcription factors were identified in the ab initio predicted genic sequences. Different machine learning algorithms were tested for classifying pre-miRNA (precursor microRNA) coding sequences, identifying Support Vector Machine (SVM) as the best performing classifier. SVM classification predicted 600 putative pre-miRNA coding loci. Repetitive DNA content of the assembly was also characterized. The first draft assembly of the quince genome produced in this work would constitute a foundation for functional genomic research in quince toward dissecting the genetic basis of important traits and performing genomics-assisted breeding.

A barcode-DNA analysis method for the identification of plant oil adulteration in milk and dairy products.

In the present work, a barcode-DNA analysis method is described for the detection of plant oil adulteration in milk and dairy products. The method relies on the fact that plant DNA should not be present in readily detectable amounts in a dairy product unless it contains undeclared plant material. Thus, a universal plant barcode is chosen as the target to be amplified from dairy samples. Accordingly, barcode PCR-CE (PCR-capillary electrophoresis) assays are described, which do not require preliminary information on the species source of the adulterant oil type. Two PCR-CE assays, one operating on the plastid trnL (UAA) intron and the other targeting its inner P6 loop in nested format, were shown to detect corn, soybean, rapeseed and sunflower oils in clarified butter, milk and yogurt. Both barcodes are robustly amplified with extremely conserved primers. While the intron provides the species discrimination ability, the P6 loop provides superior detection sensitivity.

Barcode DNA length polymorphisms vs fatty acid profiling for adulteration detection in olive oil.

The aim of this study was to compare the performance of a DNA-barcode assay with fatty acid profile analysis to authenticate the botanical origin of olive oil. To achieve this aim, we performed a PCR-capillary electrophoresis (PCR-CE) approach on olive oil: seed oil blends using the plastid trnL (UAA) intron barcode. In parallel to genomic analysis, we subjected the samples to gas chromatography analysis of fatty acid composition. While the PCR-CE assay proved equally efficient as gas chromatography analysis in detecting adulteration with soybean, palm, rapeseed, sunflower, sesame, cottonseed and peanut oils, it was superior to the widely utilized analytical chemistry approach in revealing the adulterant species and detecting small quantities of corn and safflower oils in olive oil. Moreover, the DNA-based test correctly identified all tested olive oil: hazelnut oil blends whereas it was not feasible to detect hazelnut oil adulteration through fatty acid profile analysis. Thus, the present research has shown the feasibility of a PCR-CE barcode assay to detect adulteration in olive oil.

Authentication of Botanical Origin in Herbal Teas by Plastid Noncoding DNA Length Polymorphisms.

The aim of this study was to develop a DNA barcode assay to authenticate the botanical origin of herbal teas. To reach this aim, we tested the efficiency of a PCR-capillary electrophoresis (PCR-CE) approach on commercial herbal tea samples using two noncoding plastid barcodes, the trnL intron and the intergenic spacer between trnL and trnF. Barcode DNA length polymorphisms proved successful in authenticating the species origin of herbal teas. We verified the validity of our approach by sequencing species-specific barcode amplicons from herbal tea samples. Moreover, we displayed the utility of PCR-CE assays coupled with sequencing to identify the origin of undeclared plant material in herbal tea samples. The PCR-CE assays proposed in this work can be applied as routine tests for the verification of botanical origin in herbal teas and can be extended to authenticate all types of herbal foodstuffs.

Genomic Simple Sequence Repeat Markers Reveal Patterns of Genetic Relatedness and Diversity in Sesame.

Sesame (Sesamum indicum L. syn. Sesamum orientale L.) is an orphan crop species with most molecular genetic research work done in the last decade. In this study, we used a pyrosequencing approach for the development of genomic simple-sequence repeat (SSR) markers in sesame. Our approach proved successful in identifying 19,816 nonredundant SSRs, 5727 of which were identified in a contig assembly that covers 19.29% of the sesame genome. Mononucleotide repeats were the most abundant SSR type identified in the sesame genome (48.5% of all SSRs), followed by dinucleotide SSRs (45.0%). Adenine-thymine-rich motifs were predominant, representing 81.7, 51.7, 66.5, and 22.1% of the mononucleotide, dinucleotide, trinucleotide, and tetranucleotide SSRs, respectively. As a result of this work, we introduce 933 experimentally validated sesame specific markers, 849 of which are also applicable in Sesamum mulayanum (syn. Sesamum orientale var. malabaricum Nar.), the wild progenitor of cultivated sesame. Using a subset of the newly identified SSR markers, we analyzed molecular genetic diversity and population structure of a collection of world accessions. Results of the two analyses almost overlapped and suggested correlation between genetic similarity and geographical proximity. Indeed, a pattern of gene flow among sesame diversity centers was apparent, with levels of variability in some regions similar to that seen in the domestication origin of the crop. Taken together with the high rate of genomic marker transferability detected between S. indicum and S. mulayanum, our results represent additional molecular genetic evidence for designating the two taxa as cultivated and wild forms of the same species.