Journal: Nature biotechnology

Article Title: High-throughput microbial culturomics using automation and machine learning.

doi: 10.1038/s41587-023-01674-2

Figure Lengend Snippet: Fig. 1 | A data-driven microbial isolation strategy using phenotypic and morphologic features. a, Framework of phenotype and morphology-driven strain isolation and data collection of the human gut microbiome. Human fecal samples were plated and cultured under different antibiotics selection and morphologically diverse colonies were then isolated, biobanked and analyzed by downstream sequencing. b, Setup of the automated anaerobic microbial isolation and cultivation system CAMII. c, Illustration of morphology-guided colony isolation on CAMII. Colonies grown on plates are imaged under trans- and epi-illumination and subjected to contour segmentation and morphologic features extraction. Data are analyzed by PCA to identify the set of most morphologically diverse colonies that are then isolated by an integrated colony picker. d, Illustration of diverse colony morphology on plates. Colony size and shape features were extracted from transilluminated images, and colony

Article Snippet: Gel-purified libraries were quantified by Qubit dsDNA HS assay (Thermo Fisher Scientific, Q32851) and sequenced on Illumina MiSeq platform (reagent kits: v2 300-cycles, paired-end mode) at 8 pM loading concentration with 20% PhiX spike-in (Illumina, FC-110-3001) along with custom sequencing primers spiked into Miseq reagent cartridge (6 μl of 100 μM stock; well 12: 16SV4_read1, well 13: 16SV4_index1, well 14: 16SV4_read2) following the manufacturer’s instructions.

Techniques: Isolation, Cell Culture, Selection, Sequencing, Extraction, Colony Assay

Journal: Scientific reports

Article Title: Genotyped functional screening of soluble Fab clones enables in-depth analysis of mutation effects.

doi: 10.1038/s41598-023-40241-2

Figure Lengend Snippet: Figure 1. Screening strategy illustrated. Single clones are first inoculated to well-barcoding PCR reaction, and then to corresponding coordinates on expression plate. Variable domains of all clones are amplified with custom well index primers, followed with pooling and purifying all 96 samples in single batch. In plate-barcoding PCR, pooled samples from different plates are tagged with TruSeq index adapters. Samples are again pooled, batch- purified and prepared for sequencing on MiSeq. In parallel (right), functional screening of single Fab clones is carried out. Resulting immunoassay data is then combined with NGS data, truly unique clones identified and best candidates selected for more thorough affinity screening.

Article Snippet: TruSeq Custom Amplicon kit primers (Illumina inc., USA) were used as outer indexes for plate barcoding.

Techniques: Clone Assay, Expressing, Amplification, Purification, Sequencing, Functional Assay