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Racially and ethnically minoritized populations have been historically excluded and underrepresented in research. This paper will describe best practices in multicultural and multilingual awareness-raising strategies used by the Recruitment Innovation Center to increase minoritized enrollment into clinical trials. The Passive Immunity Trial for Our Nation will be used as a primary example to highlight real-world application of these methods to raise awareness, engage community partners, and recruit diverse study participants.
The ability to synthesize capped RNA transcripts in vitro using
bacteriophage polymerases has been of considerable value in
a variety of applications. However, Pasquinelli et al.
[RNA (1995) 1:957–967] found
that one-third to one-half of the caps are incorporated in the
reverse orientation, that is, with the m7G moiety of
m7GpppG linked by a 3′-5′ phosphodiester
bond to the first nucleotide residue of the RNA chain. Such reverse
caps are unlikely to be recognized by eIF4E, based on previous studies,
and thus complicate any comparison of the translational efficiencies
of in vitro-synthesized mRNAs. We therefore designed two novel cap
P3-guanosine-5′ triphosphate and
P3-guanosine-5′ triphosphate, that are, theoretically,
incapable of being incorporated in the reverse orientation. The key reactions of pyrophosphate bond formation were achieved in anhydrous dimethylformamide
solutions employing the catalytic properties of zinc salts.
Structures were proven by 1H NMR. Transcripts produced with
SP6 polymerase using “anti-reverse” cap analogs (ARCAs) were
of the predicted length and indistinguishable in size and homogeneity from
those produced with m7GpppG or GpppG. Analysis of the transcripts
with RNase T2 and tobacco acid pyrophosphatase indicated that reverse caps
were formed with m7GpppG but not with ARCAs. Both of the ARCAs
inhibited cell-free translation with a KI
similar to that of m7GpppG. Finally, the translational efficiency
of ARCA-capped transcripts in a rabbit reticulocyte lysate was 2.3- to
2.6-fold higher than that of m7GpppG-capped transcripts. This
suggests the presence of reverse caps in conventional in vitro-synthesized
mRNAs reduces their translational efficiency.