Journal: bioRxiv

Article Title: Evaluating the reliability of tools for mRNA annotation and IRES studies

doi: 10.64898/2026.03.29.707813

Figure Lengend Snippet: (A) A-cap / stemloop IRES reporter RNA carrying the negative control “scramble 1” sequence was mixed into a polysome lysate made from HEK293T cells for five minutes on ice and loaded on a sucrose gradient. (B) The sucrose gradient was fractionated and cDNA was prepared from groups of fractions as shown (numbered 1 -6). qRT-PCR was performed to quantify the exogenous “scramble 1” RNA across the fractions, as compared to endogenous Gapdh mRNA. The “scramble 1” transcript migrated into polysomal fractions 4-6, even though it is not translated .

Article Snippet: The ARCA m 7 G-cap analog (New England Biolabs; S1411) was used at a 4:1 ratio to GFP for transcription of T7-nluc constructs, while an A-cap analog (New England Biolabs S1406), was used to generate T7HP-nluc IRES reporter RNAs.

Techniques: Negative Control, Sequencing, Quantitative RT-PCR