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. 2022 Nov 8;11:e82709. doi: 10.7554/eLife.82709

Figure 1. Bicistronic gene structure of the smORFs sloth1 and sloth2.

(A) Bicistronic gene model for sloth1 and sloth2. Zoom in shows intervening sequence (GCAAA) between sloth1 stop codon and sloth2 start codon. (B) Comparison of protein structure, amino acid length size, and amino acid percent identity between Drosophila and Human orthologs. Shaded rectangle indicates predicted transmembrane (TM) domain. (C) Phylogenetic tree of sloth1 and sloth2 orthologs in representative eukaryotic species. Linked gene structure (candidate bicistronic transcript or adjacent separate transcripts) is indicated by a black line connecting red and blue squares. (D) Plasmid reporter structure of pMT-sloth1-Rluc and derivatives. Kozak sequences upstream of start codon are underlined. Mutations indicated with shaded grey box. pMT = Metallothionein promoter. RLuc = Renilla Luciferase. (E) Quantification of RLuc luminescence/Firefly Luciferase, normalized to pMT-sloth1-Rluc, for each construct. Significance of mutant plasmid luminescence was calculated with a T-Test comparing to pMT-sloth1-Rluc. Error bars are mean with SEM. **** p≤0.0001. N=4 biological replicates.

Figure 1.

Figure 1—figure supplement 1. Related to Figure 1.

Figure 1—figure supplement 1.

(A) Comparison of gene and transcript structure of the sloth1 and sloth2 open-reading frames. A common primer pair is used to distinguish genomic from cDNA (transcript) template by PCR. Sequence of sloth1-2 genomic and sloth1-2 transcript region provided. (B) DNA gel image of PCR fragments amplified from indicated template samples. Predicted spliced transcript containing both sloth1 and sloth2 open-reading frames is amplified from cDNA generated from adult flies, 3rd instar larvae, and S2R+ cells.