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. Author manuscript; available in PMC: 2021 Aug 27.
Published in final edited form as: Nature. 2020 May 20;582(7811):283–288. doi: 10.1038/s41586-020-2323-8

Figure 2. pUG tails convert inert RNA fragments into agents of gene silencing.

Figure 2.

a-c, To control for potential dsRNA contamination in in vitro transcription reactions, RNAs were injected into rde-1(ne219) mutants, which cannot respond to dsRNA3. a, Fluorescence micrographs showing −1 to −3 oocytes of adult progeny of rde-1(ne219); gfp::h2b animals injected in the germline with in vitro transcribed RNAs consisting of the first 369nt of gfp mRNA with the indicated 3’ terminal repeats. Mean % progeny with gfp::h2b silenced is indicated ± standard deviation (s.d.). # of injected animals (n) = 3 (no injection); 9 [gfp dsRNA, (AU)18]; 10 [no tail, (GC)18, (AC)18]; and 16 [(UG)18]. b-c, oma-1(zu405ts) animals lay arrested embryos at 20°C unless oma-1(zu405ts) is silenced14. Adult rde-1(ne219); oma-1(zu405ts) animals were injected in the germline with in vitro transcribed RNAs consisting of the first 541nt of oma-1 mRNA with b, indicated 3’ terminal repeats or c, varying 3’ pUG tail lengths; different 3’ UG repeat sequences; or with (UG)18 on the 3’ end, 5’ end or in the middle of the oma-1 mRNA. For all oma-1 pUG RNA injection data, each point represents % hatched embryos laid by 5 progeny derived from one injected animal at 20°C (see Methods). Error bars: s.d. of the mean. Insets: injected RNAs run on 2% agarose gel to assess RNA integrity. For b, n=6 (no injection); 10 (oma-1 dsRNA, no tail); 12 [(UG)18]; 9 [(GC)18]; and 8 [(AU)18, (AC)18]. For c, n=9 [no injection #1, (U18G18), (UUGG)9]; 10 [(UG)0, 1, 8, 14, scrambled UGs, 3’ (UG)18, internal (UG)18]; 8 [(UG)5, 5’ (UG)18]; 12 [(UG)18 #1]; 5 [(UG)18 #2]; 11 [(UG)40]; 6 (no injection #2) and 3 (no injection #3).