# US 11,898,144 B2 · Synthetic near-threshold translational repressors

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- Number: US 11,898,144 B2
- Document ID: US11898144B2
- Status: Granted
- Kind: B2
- Application number: 17/155,387
- Filed: 2021-01-22
- Published: 2021-05-20
- Granted: 2024-02-13
- Earliest claimed: 2016-08-01
- Assignees: Arizona Board of Regents on behalf of Arizona State University
- Applicants: Arizona Board of Regents on behalf of Arizona State University
- Inventors: Alexander Green; Yu Zhou
- Web page: https://patentgrep.com/patent/US11898144B2

## CPC

- C12N 15/113: Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; {Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing (when used in plants C12N15/8218)}
- C12N 15/67: General methods for enhancing the expression
- H03K 19/20: characterised by logic function, e.g. AND, OR, NOR, NOT circuits (H03K19/003 - H03K19/01 take precedence)
- C12N 2310/122: Hairpin
- C12N 2310/531: Stem-loop; Hairpin
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- H03K: PULSE TECHNIQUE (measuring pulse characteristics G01R; modulating sinusoidal oscillations with pulses H03C; transmission of digital information H04L; discriminator circuits detecting phase difference between two signals by counting or integrating cycles of oscillation H03D3/04; automatic control, starting, synchronisation or stabilisation of generators of electronic oscillations or pulses where the type of generator is irrelevant or unspecified H03L; coding, decoding or code conversion, in general H03M)

## Abstract

Provided herein are synthetic nucleic acid molecules and methods of using such synthetic nucleic acid molecules for strong repression of target gene expression. In particular, provided herein are methods for altering expression of a protein in a cell, where the method comprises introducing into a cell a protein coding sequence operably linked to a near-threshold translational repressor having first and second trigger recognition sequences that are fully or partially complementary to a repressing trigger RNA; and introducing into a cell the repressing trigger RNA.

## Claims

### Claim 1

A synthetic nucleic acid molecule comprising a NOT-OR (NOR) logic circuit, the NOR logic circuit comprising:
at least one input RNA sensing hairpin module comprising:
an input RNA binding domain; and
a loop domain;
wherein the input RNA binding domain is complementary to an input RNA; and wherein the loop domain comprises first and second trigger RNA sequences;
a near-threshold translational repressor (NeaTTR) hairpin module comprising:
a loop-forming region comprising a ribosomal binding site (RBS); and
a stem-forming region comprising a start codon; and
first and second trigger recognition sequences located 5′ and 3′ to the NeaTTR hairpin module, respectively;
wherein the first and second trigger recognition sequences are complementary to the first and second trigger RNA sequences, respectively, of the input RNA sending hairpin module.

### Claim 2 (depends on 1)

The synthetic nucleic acid molecule of claim 1, wherein the NeaTTR hairpin module is operably linked to a reporter element.

### Claim 3 (depends on 2)

The synthetic nucleic acid molecule of claim 2, wherein the reporter element is GFP or lacZ.

### Claim 4 (depends on 1)

The synthetic nucleic acid molecule of claim 1, wherein the NOR logic circuit comprises two RNA sensing hairpin modules.

### Claim 5 (depends on 1)

The synthetic nucleic acid molecule of claim 1, wherein the NOR logic circuit comprises three RNA sensing hairpin modules.

### Claim 6 (depends on 1)

The synthetic nucleic acid molecule of claim 1, wherein the NOR logic circuit comprises four or more RNA sensing hairpin modules.