Single-Molecule FISH Probe Labeling

smFISH probes (32 probes per gene) were designed using the Stellaris Probe Designer (Biosearch Technologies) and ordered as unlabelled DNA oligos. Labelling was done according to Gaspar et al, 2017 (link) (Gaspar et al, 2017 (link)). Briefly, unlabelled ddUTP was conjugated to an ATTO dye NHS ester (ATTO565 or ATTO633, Atto-tec), then the labelled ddUTP was added to the 3’-end of each probe with terminal deoxynucleotidyl transferase. Probes were purified by ethanol precipitation.

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Samuels T.J., Gui J., Gebert D, & Karam Teixeira F. (2024). Two distinct waves of transcriptome and translatome changes drive Drosophila germline stem cell differentiation. The EMBO Journal, 43(8), 1591-1617.

Publication 2024

Stellaris Probe Designer ATTO565 ATTO633

Corresponding Organization : University of Cambridge

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Variable analysis

independent variables

Labelling method: Conjugation of unlabelled ddUTP to an ATTO dye NHS ester (ATTO565 or ATTO633), followed by addition of the labelled ddUTP to the 3'-end of each probe with terminal deoxynucleotidyl transferase

dependent variables

Not explicitly mentioned

control variables

Probe design: 32 probes per gene, designed using the Stellaris Probe Designer (Biosearch Technologies)
Probe purification: Ethanol precipitation

controls

Positive control: Not mentioned
Negative control: Not mentioned

Annotations

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