Journal: STAR Protocols

Article Title: Protocol to track single-cell-derived clones using DNA barcoding combined with single-cell RNA sequencing

doi: 10.1016/j.xpro.2025.104229

Figure Lengend Snippet: Semi-random barcode design Schematic of semi-random barcode oligonucleotide design with restriction sites used for insertion and a barcode ID tag.

Article Snippet: Package lentiviruses with HEK293T cells and the barcode library plasmid DNA using the Lipofectamine 3000 kit (Invitrogen) as per manufacturer’s instructions. a.

Techniques:

Journal: bioRxiv

Article Title: Distinct alpha-synuclein strains derived from Parkinson’s disease patient tissues trigger differential inclusion pathology in a novel biosensor cell model

doi: 10.1101/2025.04.01.646513

Figure Lengend Snippet: ( A ) Experimental design showing the timelines for generation of induced neurons from U251 FRET-based biosensor cells by reprogramming with lentiviral transduction of NEUROD1, and neuronal differentiation followed by application of skin- or brain-amplified αSyn strains with lipofectamine. ( B ) Representative confocal images showing the expression of TUJ1 and mature neuronal markers NeuN and VGLUT1, but not PROX1 at DPI 48. White arrows indicate the expanded inset image showing the subcellular localization of neuronal markers. Scale bars: 20 µm and 1µm. ( C ) A representative voltage clamp recording at -80 mV from a NEUROD1 positive cell at DPI 48 revealing large sodium currents in response to 50, 60 and 70 mV steps (300 ms); scale: 1nA/100ms. The sodium current in response to the 50 mV step is expanded and highlighted in red. Below, voltage traces from a NEUROD1 positive cell at DPI 48 in response to current injections (10, 30, 50 pA; 500 ms) revealing action potentials; scale 20 mV/100 ms. The pie chart highlights the number of NEUROD1 positive cells with action potentials. ( D-G ) Representative confocal images showing intraneuronal p-αSyn inclusions that were mostly localized in the soma. White squares indicate expanded image on the right showing the morphology and subcellular localization of p-αSyn inclusions as indicated by white arrows. Scale bars: 50µm and 3µm. ( H, I, J ) Quantification of p-αSyn inclusion fold change (H), p-αSyn inclusion lengths (I), and fold change of neuronal cell bodies (J) at DPA 14. Quantification of data are presented as mean ± SEM, and each dot corresponds to data analyzed from 180-700 neurons in a biological replicate. N=10-12 from the skin-amplified strains of 2 cases and N=11-12 from the brain-amplified strains of 1 case, N=10-12 from rPFF group, and N=11-12 from monomer group from 2 independent differentiations. Statistical analysis was performed by ordinary one-way ANOVA with Tukey’s multiple comparison test or Brown-Forsythe and Welch ANOVA with Dunnet’s T3 multiple comparison test (H). A p value <0.05 was considered significant. Significant differences are indicated by *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.

Article Snippet: EF1a_NEUROD1_P2A_Hygro_Barcode (Addgene plasmid # 120466) was a gift from Prashant Mali [ ].

Techniques: Transduction, Amplification, Expressing, Comparison