Journal: bioRxiv

Article Title: An updated C. elegans body muscle transcriptome for studies in muscle formation and function

doi: 10.1101/2022.04.12.488068

Figure Lengend Snippet: A) Transgenic worms expressing mCherry fluorochrome fused to histone H2B ( his-58 ) are (1) homogenized, (2) then subjected to several rounds of mechanical filtration. (3) The resultant nuclei are then collected and (4) subjected to FACS sorting. At the completion of the sorting step, (5) the mCherry-positive nuclei were incubated into TRIzol reagent and the recovered RNA is sequenced. B) Top panel: The strain BN452 ubiquitously expresses the mCherry:: his-58 transgene. Bottom Panel: mCherry positive nuclei recovered after Step 2 described in Panel A. (Red: mCherry, Blue: DAPI). C) Western blot of nuclear and cytoplasmic fractions from BN452 worms blotted with α-RFP and α-GAPDH antibodies. D) Left Panel: nuclear FACS profile from N2 and BN452 nuclei. The red dots show the sorted mCherry-positive population. Right Panel: Pie chart showing the percentage of BN452 sorted mCherry-positive nuclei.

Article Snippet: Two replicates of resuspended nuclei samples for the body muscle tissue were FACS isolated into TRIzol solution using a BD Biosciences FACSAria III cell sorter with a 70 µm nozzle, and a 560 nm laser with a temperature-controlled tube holder at 4°C.

Techniques: Transgenic Assay, Expressing, Filtration, Incubation, Western Blot