๐ฏ๐ I've been away for a while. But here:
I wrote a blog post on "Imaging mRNA modifications in situ"
Featuring: m6AISH-PLA, ARPLA (on #glycoRNA) & DART-FISH.
#smFISH #m6A #RNA #mRNA #RNAmodifications #microscopy #imaging #singlemolecule
https://greenfluorescentblog.wordpress.com/2023/10/31/imaging-mrna-modifications-in-situ/
Wow! ๐ฅ ๐ฅ ๐ฅ ๐ฅ
This is so cool!
In this paper,
"Spatially resolved single-cell translatomics at molecular resolution", they used a tri-probe FISH system combined with in situ amplification and in situ sequencing to detect multiplex translating mRNAs at sub-cellular resolution in cell culture (981 genes) and mouse brain tissue slices (5413 genes!).
Wow!
looks like super awesome method from Long Cai lab
"High-resolution spatial multi-omics reveals cell-type specific nuclear compartments"
They did mRNA #seqFISH+ (1200 genes, 17K introns), DNA seqFISH+ (100K loci) and #immunofluorescence (up to 65 antibodies) all in the same experiment!
200+ rounds of #imaging! That's incredible.
#mRNA #mRNALocalization #smFISH #DNA #3D_genome
#transcription #microscopy
"HT-smFISH: a cost-effective and flexible workflow for high-throughput single-molecule RNA imaging"
In HT-smFISH, multiple RNA probes are generated by parallel in vitro transcription from a large pool of unlabeled oligonucleotides. This reduces costs per targeted RNA compared with many smFISH methods and is easily scalable and flexible in design.
I tried the original protocol a few years ago and didn't work well. inconsistent labeling (some good, some poor) and wasn't *that* cheap.
but maybe time to re-visit this protocol with this improved version:
"Improved enzymatic labeling of fluorescent in situ hybridization probes applied to the visualization of retained introns in cells"
@biorxivpreprint
RS-FISH is a user-friendly software for accurate spot detection that is applicable to smFISH experiments, spatial transcriptomics, and spatial genomics. The approach enables fast spot detection in even very large volumetric datasets.
"RS-FISH: precise, interactive, fast, and scalable FISH spot detection"
New, user-friendly, FISH spot detection software; runs as a #Fiji plugin.
From Stephen Preibisch lab with @successprocess and Kyle Harrington
Out in @naturemethods
#mRNA #fluorescence #smFISH #SingleMolecule #microscopy #imaging
RS-FISH is a user-friendly software for accurate spot detection that is applicable to smFISH experiments, spatial transcriptomics, and spatial genomics. The approach enables fast spot detection in even very large volumetric datasets.
Gal Haimovich ๐ท๐ฌโฃ๐ณ๏ธโ๐๐ณ๏ธโโง