Journal: PLoS Genetics

Article Title: Order of removal of conventional and nonconventional introns from nuclear transcripts of Euglena gracilis

doi: 10.1371/journal.pgen.1007761

Figure Lengend Snippet: Categories of sequencing reads inferred from the PacBio NGS sequencing. (A) Categories of sequencing reads deriving from the gapC amplicon, (B) i6-tubA amplicon and (C) i5-tubA amplicon, respectively. Number and percentage of particular types of reads are given. The distribution of introns in all groups of reads is depicted. Lines represent the exons while circles represent the introns: black circles: conventional introns; white: nonconventional ones (not to scale). Introns are labelled i1 –i6. In addition, introns that exhibit intermediate features are marked with an asterisk. Double asterisk indicates the introns to which the reverse primer was targeted, thus not considered in the analysis of amplicons (introns to which primers annealed were inevitably present in the amplification products).

Article Snippet: The amplicons obtained as a result of RT-PCR reactions for the gapC and tubA genes were sequenced commercially in the Museum and Institute of Zoology of the Polish Academy of Science, Warsaw, Poland, on the PacBio RS II (Pacific Biosciences) instrument.

Techniques: Sequencing, Next-Generation Sequencing, Amplification

Journal: BMC Genomics

Article Title: Haplotyping and copy number estimation of the highly polymorphic human beta-defensin locus on 8p23 by 454 amplicon sequencing

doi: 10.1186/1471-2164-11-252

Figure Lengend Snippet: Human defensin gene clusters at 8p23.1 (NCBI build 36, hg18) and location of the amplicons analyzed by 454 (F1-8) and cloning/Sanger (S1-2) sequencing within the proximal DEFB (DEF cluster b2) .

Article Snippet: The pools were diluted and subjected to emulsion PCR following the FLX emPCR protocol for amplicons (Roche Diagnostics, December 2007) using both emPCR kits II (primer A) and III (primer B) and sequenced on a GS FLX (Roche Diagnostics) by both primers on 1 lane/pool of a 16-lane gasket on a 70 × 75 FLX picotiterplate.

Techniques: Clone Assay, Sequencing

Journal: Scientific Reports

Article Title: Rapid tumor induction in zebrafish by TALEN-mediated somatic inactivation of the retinoblastoma1 tumor suppressor rb1

doi: 10.1038/srep13745

Figure Lengend Snippet: rb1 mutant alleles and allele frequencies in somatic, germline, and tumor tissues from adult genetic mosaic zebrafish. ( a ) 14 different indel alleles in rb1 exon 2 were identified in retina, muscle, and germline genomic DNA from two adults. Mutant alleles represented 33% of cloned amplicons. In tumor tissue 8 unique alleles were identified, representing 87% of cloned amplicons. ( b ) Two frameshift alleles that delete 11 bp and 19 bp in rb1 exon 3 were detected in two adults. 10% of sequenced clones were mutant in retina, muscle or germline tissue. In tumor tissue 68% of sequenced clones were mutant alleles. Red arrows mark TALEN cut site.

Article Snippet: Amplicons were TOPO cloned (Invitrogen), and 50–100 individual clones sequenced per sample.

Techniques: Mutagenesis, Clone Assay

Journal: Limnology and oceanography, methods / ASLO

Article Title: Molecular detection of harmful algal blooms (HABs) using locked nucleic acids and bead array technology

doi: 10.4319/lom.2010.8.269

Figure Lengend Snippet: Analytic detection limits of chemically and enzymatically labeled amplicons. The amplicons were serially diluted and hybridized for 1.5 h at 53°C. The error bars represent the standard deviation. The samples were run in duplicate, and the experiments

Article Snippet: Chemically labeled amplicons (Mirus Label IT amplicons): For chemically labeled amplicons, illustrates the detection levels of the assay conducted at 53°C.

Techniques: Labeling, Standard Deviation

Journal: Limnology and oceanography, methods / ASLO

Article Title: Molecular detection of harmful algal blooms (HABs) using locked nucleic acids and bead array technology

doi: 10.4319/lom.2010.8.269

Figure Lengend Snippet: Performance of species-specific probes with field samples. Hybridization assay conditions used 15 μL chemical-labeled IT amplicon. The amplicons were generated using 40 cycles, and amplification was done with the universal primer set D2C and DIRF.

Article Snippet: Chemically labeled amplicons (Mirus Label IT amplicons): For chemically labeled amplicons, illustrates the detection levels of the assay conducted at 53°C.

Techniques: Hybridization, Labeling, Amplification, Generated

Journal: Limnology and oceanography, methods / ASLO

Article Title: Molecular detection of harmful algal blooms (HABs) using locked nucleic acids and bead array technology

doi: 10.4319/lom.2010.8.269

Figure Lengend Snippet: Comparison of Mirus Label IT and 5′ end labeled amplicons. Fifty nanograms of the labeled template was hybridized with different sets of K. brevis (Pkb, Pkb2, PKB658, Pkb2lna, PKB658lna) and K. mikimotoi probes (Pkmiklna and Pkmiklna2). Hybridization

Article Snippet: Chemically labeled amplicons (Mirus Label IT amplicons): For chemically labeled amplicons, illustrates the detection levels of the assay conducted at 53°C.

Techniques: Labeling, Hybridization

Journal: Limnology and oceanography, methods / ASLO

Article Title: Molecular detection of harmful algal blooms (HABs) using locked nucleic acids and bead array technology

doi: 10.4319/lom.2010.8.269

Figure Lengend Snippet: Specificity of probes with Mirus Label IT D1/D2 amplicons. Probes were challenged with closely and non–closely related algal species. The hybridization was carried out for 1.5 h at 53°C and used 33 ng amplicon per well. The samples were

Article Snippet: Chemically labeled amplicons (Mirus Label IT amplicons): For chemically labeled amplicons, illustrates the detection levels of the assay conducted at 53°C.

Techniques: Hybridization, Amplification

Journal: Scientific Reports

Article Title: Quantifying perinatal transmission of Hepatitis B viral quasispecies by tag linkage deep sequencing

doi: 10.1038/s41598-017-10591-9

Figure Lengend Snippet: Tag-linkage sequencing of HBV RT/S region. ( a ) A schematic presentation of the HBV viral genome region (polymerase gene) that was processed and reconstructed using the tag-linkage method. 836 bp of polymerase gene (blue box) was recovered, including the majority of the reverse transcriptase domain (overlapping S gene) and part of the RNase H gene. ( b ) A schematic presentation of the experiment flow for tag linkage deep sequencing. 1) HBV sequences were amplified from viral DNA extracted from patient plasma using primers targeting conserved region. Tags were added onto each molecule for sequencing error correction and haplotype reconstruction. 2) Different length of sequences (covering different number of non-overlapping small amplicons) were amplified from ~10,000 molecules. 3) Amplified sequences were ligated. 4) Small amplicons were further amplified for sequencing.

Article Snippet: Finally, sequencing adaptors were added onto amplicons for Illumina sequencing.

Techniques: Sequencing, Flow Cytometry, Amplification

Journal: Scientific Reports

Article Title: A comparative assessment of conventional and molecular methods, including MinION nanopore sequencing, for surveying water quality

doi: 10.1038/s41598-019-51997-x

Figure Lengend Snippet: Results of the experimentally determined composition of the MOCK Community (MC) at ( a ) family and ( b ) genus level with MinION and Illumina NGS techniques. The results are based on 16S rRNA amplicon sequencing. Experimentally determined 16S rRNA gene percentage abundance are compared against the actual composition of the MC as reported by the supplier (i.e. Zymo Research). Data points are an average of duplicate samples for Illumina and triplicate samples for MinION sequencing. Error bars indicate the standard deviation.

Article Snippet: The amplicon data from Illumina were processed using an open source software package: Quantitative Insights Into Microbial Ecology, QIIME 2 ( https://qiime2.org/ ).

Techniques: Next-Generation Sequencing, Amplification, Sequencing, Standard Deviation

Journal: Neurology: Genetics

Article Title: Brain somatic mutations in SLC35A2 cause intractable epilepsy with aberrant N-glycosylation

doi: 10.1212/NXG.0000000000000294

Figure Lengend Snippet: Representative radiologic and pathologic images of patients with brain somatic mutations in SLC35A2 (A) Preoperative and postoperative brain MRI T2-weighted images from patients EPI219 and LGS150 with brain somatic mutations in SLC25A2 . These T2-weighted images demonstrate no remarkable findings in the brain parenchyma, including the temporal lobe. Yellow arrowhead: putative regions of epileptic focus. (B) Histopathologic images from H E staining and immunohistochemical (IHC) staining from EPI219 (upper panels) and LGS150 (lower panels) brain tissues. Black arrowheads: scattered neuron in white matter. Scale bars, 40 μm in H E staining and 200 μm in IHC staining for NeuN, a neuronal marker (C) Capture image from integrative genomic viewer (IGV) (upper panels), showing the results of site-specific amplicon sequencing. Schematic tables (lower panels) showing the number of sequence reads counted as mutated or reference sequences, as well as the VAFs of mutated alleles. Mut: mutation, Ref: reference.

Article Snippet: Briefly, this method calculates the discrepancy between expected and observed amounts of mismatches in amplicon-based, Illumina platform data sets (up to 10,000X) in which 2 independent blood samples with known single nucleotide polymorphisms (SNPs) were mixed to mimic somatic mutations with 4 different VAFs: 0.5%, 1%, 5%, and 10%.

Techniques: Magnetic Resonance Imaging, Staining, Immunohistochemistry, Marker, Amplification, Sequencing, Mutagenesis

Journal: Genome Biology and Evolution

Article Title: Attenuated Virulence and Genomic Reductive Evolution in the Entomopathogenic Bacterial Symbiont Species, Xenorhabdus poinarii

doi: 10.1093/gbe/evu119

Figure Lengend Snippet: The xaxAB locus, its genomic context and its shuffling point exbD / rdgC in the X. doucetiae FRM16 ( Xd ), X. nematophila ATCC19061 ( Xn ), X. bovienii SS-2004 ( Xb ), X. poinarii G6 ( Xp_G6 ), AZ26 ( Xp_AZ26 ), NC33 ( Xp_NC33 ), SK72 ( Xp_SK72 ), and CU01 ( Xp_CU01 ) genomes. The large arrows represent individual ORFs, and the names of the genes are indicated above the arrows. Genes encoding proteins of unknown function are marked with an asterisk. Orthologous genes are indicated by arrows in the same color. Black and chequered arrows represent core-genome genes and transposase genes, respectively. The thin arrows indicate the binding sites of the primers used for PCR amplification. The vertical parallel lines indicate the end of the sequenced area and the dotted lines represent an unsequenced genomic region. The cladogram was obtained by the maximum-likelihood phylogenetic analysis of five concatenated protein-coding sequences ( recA , gyrB , dnaN , gltX , and infB ), as already described in figure 1 . The accession numbers of the sequences of the subsequent amplicons are HG934736 (strain AZ26), HG934737 (strain NC33), HG934738 (strain SK72), HG934739 and HG934740 (strain CU01).

Article Snippet: Amplicons were sequenced by MWG-Eurofins France.

Techniques: Binding Assay, Polymerase Chain Reaction, Amplification