Journal: bioRxiv

Article Title: Mosaic Ends Tagmentation (METa) assembly for extremely efficient construction of functional metagenomic libraries

doi: 10.1101/2021.02.01.429292

Figure Lengend Snippet: The general pipeline for the creation and use of functional metagenomic libraries to capture and discover genes from metagenomes. 1) Extraction of metagenomic DNA from a microbiome ( e.g. soil or fecal samples). 2) Fragmentation of metagenomic DNA to desired size range ( e.g. by sonication, restriction enzyme digestion, or tagmentation). 3) Cloning of fragments into expression vectors following size selection ( e.g. by blunt ligation or homology-based assembly. 4) Transformation en masse of vectors into an expression host ( e.g. E. coli DH10B) to create functional metagenomic library. 5) Functional selection or screen of library ( e.g. on antibiotics to select for resistance). 6) Amplification of selected inserts using vector-specific primers. 7) High-throughput sequencing of selected metagenomic amplicons ( e.g. by Illumina or PacBio technologies). 8) Annotation of sequenced amplicons to link novel genes with selected/screened function ( e.g. discovery of novel aminoglycoside acetyltransferases). Figure created in BioRender.com.

Article Snippet: For each reaction, the entire 10 μl milliQ water elution was electroporated into a 25 μl aliquot of commercial 10-beta electrocompetent E. coli DH10B cells (advertised transformation efficiency of 2×10 10 cfu/μg DNA), immediately rescued in 1 ml of 37°C SOC outgrowth medium (New England Biolabs, cat#B9020S) and incubated with shaking at 37°C for 1 hour.

Techniques: Functional Assay, Sonication, Clone Assay, Expressing, Selection, Ligation, Transformation Assay, Amplification, Plasmid Preparation, Next-Generation Sequencing

Journal: eLife

Article Title: Reporter-ChIP-nexus reveals strong contribution of the Drosophila initiator sequence to RNA polymerase pausing

doi: 10.7554/eLife.41461

Figure Lengend Snippet:

Article Snippet: For plasmid amplification and purification, the DNA was transformed into NEB 5-alpha Competent E. coli (High Efficiency) or One Shot ccdB Survival 2 T1R Competent Cells and purified with IBI High Speed Plasmid Mini Kit.

Techniques: Cell Culture, Protease Inhibitor, SYBR Green Assay, Plasmid Preparation, Multiplex Assay, Mutagenesis, Ligation, Sequencing, RNA Sequencing Assay, Recombinant, Software

Journal: Journal of Clinical Microbiology

Article Title: Rapid Detection of Hepatitis B Virus Variants Associated with Lamivudine and Adefovir Resistance by Multiplex Ligation-Dependent Probe Amplification Combined with Real-Time PCR

doi: 10.1128/JCM.02554-13

Figure Lengend Snippet: Outline of MLP-RT-PCR assay. (a) 5′ Probe and 3′ probe. (b) Hybridization of short probes to the target sequences. (c) The short probes were ligated by the DNA ligase enzyme, and the ligated products were amplified with a pair of common primers. (d) Real-time PCR was performed, and the TaqMan probe was hydrolyzed by DNA polymerase to release the fluorophore.

Article Snippet: The ligation efficiency of Taq DNA ligase (NEB, USA) was greater than that of ligase-65 (MRC-Holland, Holland, the Netherlands).

Techniques: Reverse Transcription Polymerase Chain Reaction, Hybridization, Amplification, Real-time Polymerase Chain Reaction