SELEX Screening and
Characterization of Small RNA Molecules That Specifically Bind the Reactive
Blue Dye
WANG Chen, XU Feng, JIN You-Xin and WANG De-Bao
( State Key Laboratory of Molecular Biology, Shanghai Institute of
Biochemistry, the Chinese Academy of Sciences, Shanghai 200031, China
)
Abstract A 73-base single-stranded DNA
library with a T7 promoter was chemically synthesized, in which 20 consecutive
bases were completely randomized. After PCR amplification and T7 RNA polymerase
transcription of the DNA random library, the in vitro transcribed RNA
random library was subjected to 8 successive rounds of reactive blue dye column
selection by SELEX method, resulting in a sharp increase of the percentage of
the transcribed RNA pool that could bind reactive blue dye, from less than
0.03% in the first round pool to 22.4% in the eighth-round pool. Cloning and
sequencing of the enriched RNAs pool led to three groups of RNA molecules in
terms of molecular lengths. The correlation between the binding capacities and
their corresponding secondary structures of the screened RNAs suggests that the
RNA helix is the main motif in the interaction between RNAs and reactive blue
dye. These results demonstrate that it is possible to find RNA candidates with
a given function in a shorter RNA random library so that it is easier to follow
the structure and function relationship. In addition, longer RNA sequences
could theoretically be derived from short RNA sequences through a process of
motif combination in the course of molecular evolution.
Key Words SELEX； reactive blue dye； biomacromolecule evolution； small RNA
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