double stranded barcode dna  (Thermo Fisher)

Journal: bioRxiv

Article Title: Plasticity of extrachromosomal DNA segregation during drug adaptation

doi: 10.64898/2025.12.09.693337

Figure Lengend Snippet: Schematic of the scDNA-seq experimental workflow. The target regions were amplified, followed by barcode assignment within the emulsions. The emulsions were broken, adapters were added, and libraries were prepared for NGS. b, Reference regions used for normalization are indicated by red arrows. One or two reference regions were designed for each chromosome. c, Schematic of the two types of celularl barcodes, PseuMO-Tag and ID vector.

Article Snippet: For insertion, 2 μL of 100 μM barcode oligonucleotide and 1 μL of 100 μM reverse primer (5′-TGCAGCATGCGTCTCACAACG-3′) were mixed with 25 μL of NEBNext High-Fidelity 2× PCR Master Mix (New England Biolabs) and 22 μL of water to produce double-stranded barcode DNA (Thermo Fisher Scientific).

Techniques: Amplification, Plasmid Preparation

Journal: bioRxiv

Article Title: Plasticity of extrachromosomal DNA segregation during drug adaptation

doi: 10.64898/2025.12.09.693337

Figure Lengend Snippet: Schematic of single-cell cloning with cellular barcode. After PseuMO-Tag lentiviral cellular barcodes were transduced into each cell line, single cells were isolated into 96-well plates. Each clone was cultured until it reached approximately 1–2 × 10⁵ cells and cryopreserved. Once a sufficient number of clones was established, pooled clones were analyzed using scDNA-seq. For COLO320DM, the same clonal pool was additionally analyzed with scRNA-seq, and each clone was cultured individually. After 18 additional cell divisions, scDNA-seq was performed. b, Gamma distribution fitting of copy-number distributions of MYC for each clone derived from COLO320DM, H2170, and PC-3 cells. The dataset includes 5,861 cells from 12 clones in COLO320DM; 7,118 cells from 21 clones in H2170; and 14,796 cells from 25 clones in PC-3 cells. c, Heatmap of eight parameters of MYC for each clone shown in b, sorted in descending order of the mean value. Each column was normalized by Z-score scaling. d, Scatter plot of the mean and SD values for MYC copy number showing differences among clones. e, Gamma distribution fitting of MYC copy-number distributions in each COLO320DM clone. Orange curves represent the distributions at the same time point as in Fig. 2b –d and Extended Data Fig. 2. Purple curves represent the distributions after 18 additional cell divisions. f, Density plot of MYC expression levels for each clone. g, Correlation of the mean of MYC copy number and Mean MYC expression levels (left), and SD of MYC copy number and SD of MYC expression levels (right). h, Correlation of the mean of MYC copy number and mean of MYC target expression levels.

Article Snippet: For insertion, 2 μL of 100 μM barcode oligonucleotide and 1 μL of 100 μM reverse primer (5′-TGCAGCATGCGTCTCACAACG-3′) were mixed with 25 μL of NEBNext High-Fidelity 2× PCR Master Mix (New England Biolabs) and 22 μL of water to produce double-stranded barcode DNA (Thermo Fisher Scientific).

Techniques: Cloning, Isolation, Cell Culture, Clone Assay, Derivative Assay, Expressing

Journal: Nucleic Acids Research

Article Title: STAG2 loss-of-function affects short-range genomic contacts and modulates the basal-luminal transcriptional program of bladder cancer cells

doi: 10.1093/nar/gkab864

Figure Lengend Snippet: Rewiring of chromatin looping arising from loss of STAG2 results in transcriptional changes. ( A ) Heatmap representing the statistical significance (FDR) of motif analyses of promoters of differentially expressed genes and genomic positions involved in differential DNA looping in STAG2-silenced cells. We identify three collections of motifs: #1, motifs enriched in differential interactions; #2, motifs enriched in differentially expressed genes; and #3, motifs enriched in both interactions and transcriptionally deregulated genes. ( B ) Average fold change in gene expression values (FPKM) of genes engaged by control and differential interactions overlapping promoters or ( C ) gene bodies. Boxplot notches represent the confidence interval around the median. t -test: * P < 0.05; ** P < 0.01; *** P < 0.001. ( D , E ) Hi-C contact matrices at the TNC and COL17A1 loci in control and STAG2-silenced cells. The contact matrices for the STAG2 -silenced condition are the result of averaging the matrices for sh1 and sh2. Snapshots of the ChIP-Seq tracks for STAG1 and STAG2, differential contact matrices, H3K27me3 peaks, and gene expression values (FPKM) are included. Loss of interactions overlapping the promoter of TNC and COL17A1 upon STAG2 silencing correlates with a consistent increase in gene expression. Error bars represent mean ± SEM.

Article Snippet: The ends of the DNA fragments were ligated to double stranded barcoded DNA adapters (NEXTflex ChIP-Seq Barcodes, Bioo Scientific, ref. 514120) using T4 DNA Ligase.

Techniques: Gene Expression, Control, Hi-C, ChIP-sequencing