Journal: Frontiers in Molecular Biosciences

Article Title: A systematic review of the barcoding strategy that contributes to COVID-19 diagnostics at a population level

doi: 10.3389/fmolb.2023.1141534

Figure Lengend Snippet: Schematic representation of mechanistic strategies of barcoding. (A–C) Barcodes can be introduced to a template using adaptors through direct ligation (A) , using RT- or PCR primers at the reverse transcription or PCR amplification step (B) , and using hybridizing molecular inversion probes (C) . (D) Schematic representation of the difference between “barcodes” and “sample indexes”. Barcodes aim to correct sequencing errors. For example, a misreading nucleotide, guanosine (G) can be corrected in final consensus sequences for a pool of Sample 1 (top panel). Sample indexes are used to multiplex different sequencing amplicons generated from different pools of samples (Sample 1, 2, and 3) (bottom panel). Panel (A) is modified based on in and panel (C) is modified based on in .

Article Snippet: Primer-associated approach , Sequence-based barcodes , SQK-RBK004: transposase carrying barcodes to the site of the cleavage , - , - , Whole genome , Oxford Nanopore Rapid Barcoding kit (SQK-RBK004) , SARS-CoV-2 patient samples (nasopharyngeal swab) , Oxford Nanopore , Guppy version 3.6.0; ARTIC Network bioinformatics protocol , Multiplex samples , Propose a method to sequence the whole genome of SARS-CoV-2 in a rapid and cost-efficient manner , .

Techniques: Ligation, Reverse Transcription, Amplification, Sequencing, Multiplex Assay, Generated, Modification

Journal: Frontiers in Molecular Biosciences

Article Title: A systematic review of the barcoding strategy that contributes to COVID-19 diagnostics at a population level

doi: 10.3389/fmolb.2023.1141534

Figure Lengend Snippet: Systematic comparison of barcoding strategies used in the category of molecular barcodes.

Article Snippet: Primer-associated approach , Sequence-based barcodes , SQK-RBK004: transposase carrying barcodes to the site of the cleavage , - , - , Whole genome , Oxford Nanopore Rapid Barcoding kit (SQK-RBK004) , SARS-CoV-2 patient samples (nasopharyngeal swab) , Oxford Nanopore , Guppy version 3.6.0; ARTIC Network bioinformatics protocol , Multiplex samples , Propose a method to sequence the whole genome of SARS-CoV-2 in a rapid and cost-efficient manner , .

Techniques: Comparison, Software, Sequencing, Multiplex Assay, CRISPR, Plasmid Preparation, Microarray, Binding Assay, Amplification, Extraction, Ligation, DNA Sequencing, Multiplexing, Generated, Reverse Transcription, Staining, Flow Cytometry, High Throughput Screening Assay, Inhibition, Blocking Assay, Conjugation Assay, RNA Sequencing Assay, Transmission Assay, Incubation, Diagnostic Assay, Next-Generation Sequencing, Infection

Journal: Applied and Environmental Microbiology

Article Title: Sources of Bacteria in Outdoor Air across Cities in the Midwestern United States

doi: 10.1128/aem.05498-11

Figure Lengend Snippet: FIG. 1. Most abundant bacterial groups identified using barcoded pyrosequencing at the phylum level (A) and at the order level (B). Proteobacterial groups are designated by the letters , , , and for the Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, and Deltaproteobacteria, respectively.

Article Snippet: Bacterial community composition was determined using a barcoded pyrosequencing procedure, which facilitates multiplexed sequencing of partial 16S rRNA genes.

Techniques:

Journal: bioRxiv

Article Title: Arrayed in vivo barcoding for multiplexed sequence verification of plasmid DNA and demultiplexing of pooled libraries

doi: 10.1101/2023.10.13.562064

Figure Lengend Snippet: (A) Donor cell arrays (blue) containing donor plasmids are conjugated to recipient cell arrays (gray) containing recipient plasmids. A DNA cassette from the donor plasmid is recombined into the recipient plasmid backbone to join an indexing DNA barcode with a sequence of interest. Cells from one or more plates can subsequently be pooled and prepared for sequencing. (B) In one design, a cassette with a DNA block on a donor plasmid is recombined into a recipient plasmid backbone containing a positional barcode. (C) In another design, where a whole plasmid backbone needs to be sequenced, a donor positional barcode is recombined into a recipient plasmid. In both (B) and (C), a scissor icon is an SceI cut site, HR is a region of sequence identity that mediates homologous recombination, dashed lines are homologous recombination events, and + or - icons are selection or counter-selection markers. (D) The recovery rate (percent of positions that were detected, orange) and accuracy (percent of detections that were sequence correct, blue) for the design in (B). Number of matings (x-axis) indicates the number of times the same DNA block was mated to a barcode and sequenced. Error bars indicate standard errors calculated by bootstrapping. (E) In the design illustrated in (B), donor barcodes were mated to recipient plasmids and plasmid sequences at each position were assembled de novo by sequencing pools of clones. Variation from the a priori reference expectation, including insertions, deletions, and substitutions are shown by colored dots. Successful de novo assemblies are ranked by decreasing ONT read coverage. (F) The cost of plasmid sequencing when BPS is performed at high- and low-throughput. Low-throughput assumes barcoding is performed in 96-well plates and 4,608 plasmids are sequenced at 100-400× coverage per flow cell. High-throughput assumes barcoding is performed on 384-position agar arrays and 9,216 plasmids are sequenced at 100-200× coverage per flow cell. Detailed cost assumptions are listed in Tables S1 and S2. (G) Experimental timeline for sequence verification by BPS. We assume a Minion flow cell contains 250 active pores generating data at 100 bases/second, enabling ∼500 (7kb) plasmids to be sequenced at 100x depth in 4 hours.

Article Snippet: In vitro methods have been developed to multiplex high-throughput sequencing by introducing DNA barcode “indices” via PCR ( – ) or Tn5 transposase tagmentation ( , ) (see also https://www.octant.bio/blog-posts/octopus-v3 ), enabling the Illumina or Oxford Nanopore Technology (ONT) sequencing platforms to sequence many barcoded samples at once.

Techniques: Plasmid Preparation, Sequencing, Blocking Assay, Homologous Recombination, Selection, Clone Assay, High Throughput Screening Assay