Source 1primary paper2022
Toolkit/tethered PEs
tethered PEs
Also known as: tPEs
Taxonomy: Mechanism Branch / Architecture. Workflows sit above the mechanism and technique branches rather than replacing them.
Summary
Tethered prime editors (tPEs) are prime editing constructs in which stem-loop aptamer-modified pegRNAs are tethered to Cas9 nickase. The design alters pegRNA architecture to increase prime editing efficiency and flexibility.
Usefulness & Problems
Why this is useful
tPEs are useful because they address the possibility that the 3-prime extension of pegRNAs can impair pegRNA stability or folding and thereby compromise prime editing activity. By tethering stem-loop pegRNAs to Cas9 nickase, the system is intended to improve the functional performance and design flexibility of prime editing constructs.
Source:
The resulting split pegRNA prime editors (SnPEs) maintain the PE activity and increase flexibility.
Problem solved
This tool is designed to solve a pegRNA architecture problem in prime editing, specifically the risk that the 3-prime pegRNA extension negatively affects RNA stability or folding. It also addresses the need for more flexible prime editor construct formats through tethered and related split pegRNA designs.
Problem links
Need conditional recombination or state switching
DerivedTethered prime editors (tPEs) are prime editing constructs in which stem-loop aptamer-modified pegRNAs are tethered to Cas9 nickase. The design alters pegRNA architecture to increase prime editing efficiency and flexibility.
Need controllable genome or transcript editing
DerivedTethered prime editors (tPEs) are prime editing constructs in which stem-loop aptamer-modified pegRNAs are tethered to Cas9 nickase. The design alters pegRNA architecture to increase prime editing efficiency and flexibility.
Taxonomy & Function
Primary hierarchy
Mechanism Branch
Architecture: A reusable architecture pattern for arranging parts into an engineered system.
Techniques
No technique tags yet.
Target processes
editingrecombinationImplementation Constraints
cofactor dependency: cofactor requirement unknownencoding mode: genetically encodedimplementation constraint: context specific validationoperating role: regulatorswitch architecture: split
tPEs are implemented by adding stem-loop aptamers to the 3-prime terminus of the pegRNA and tethering the resulting stem-loop PE to Cas9 nickase. The available evidence does not specify the aptamer identity, the tethering module, expression format, or delivery method.
The supplied evidence is limited to design rationale and construct strategy from a single 2022 source. No specific editing outcomes, target loci, organismal validation, or comparative efficiency measurements are provided here.
Validation
Cell-freeBacteriaMammalianMouseHumanTherapeuticIndep. Replication
Supporting Sources
Ranked Claims
The 3-prime extension of pegRNAs could negatively affect pegRNA stability or folding and compromise prime editing activity.
The 3’-extension of pegRNAs could negatively affect its stability or folding and comprise the PE activity.
The 3-prime extension of pegRNAs could negatively affect pegRNA stability or folding and compromise prime editing activity.
The 3’-extension of pegRNAs could negatively affect its stability or folding and comprise the PE activity.
The 3-prime extension of pegRNAs could negatively affect pegRNA stability or folding and compromise prime editing activity.
The 3’-extension of pegRNAs could negatively affect its stability or folding and comprise the PE activity.
The 3-prime extension of pegRNAs could negatively affect pegRNA stability or folding and compromise prime editing activity.
The 3’-extension of pegRNAs could negatively affect its stability or folding and comprise the PE activity.
The 3-prime extension of pegRNAs could negatively affect pegRNA stability or folding and compromise prime editing activity.
The 3’-extension of pegRNAs could negatively affect its stability or folding and comprise the PE activity.
The 3-prime extension of pegRNAs could negatively affect pegRNA stability or folding and compromise prime editing activity.
The 3’-extension of pegRNAs could negatively affect its stability or folding and comprise the PE activity.
The 3-prime extension of pegRNAs could negatively affect pegRNA stability or folding and compromise prime editing activity.
The 3’-extension of pegRNAs could negatively affect its stability or folding and comprise the PE activity.
Stem-loop PEs can be tethered to Cas9 nickase to produce tethered PEs.
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Stem-loop PEs can be tethered to Cas9 nickase to produce tethered PEs.
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Stem-loop PEs can be tethered to Cas9 nickase to produce tethered PEs.
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Stem-loop PEs can be tethered to Cas9 nickase to produce tethered PEs.
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Stem-loop PEs can be tethered to Cas9 nickase to produce tethered PEs.
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Stem-loop PEs can be tethered to Cas9 nickase to produce tethered PEs.
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Stem-loop PEs can be tethered to Cas9 nickase to produce tethered PEs.
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Stem-loop PEs were generated by adding stem-loop aptamers at the 3-prime terminal of pegRNA.
Here we generated stem-loop PEs (sPEs) by adding stem-loop aptamers at the 3’-terminal of pegRNA
Stem-loop PEs were generated by adding stem-loop aptamers at the 3-prime terminal of pegRNA.
Here we generated stem-loop PEs (sPEs) by adding stem-loop aptamers at the 3’-terminal of pegRNA
Stem-loop PEs were generated by adding stem-loop aptamers at the 3-prime terminal of pegRNA.
Here we generated stem-loop PEs (sPEs) by adding stem-loop aptamers at the 3’-terminal of pegRNA
Stem-loop PEs were generated by adding stem-loop aptamers at the 3-prime terminal of pegRNA.
Here we generated stem-loop PEs (sPEs) by adding stem-loop aptamers at the 3’-terminal of pegRNA
Stem-loop PEs were generated by adding stem-loop aptamers at the 3-prime terminal of pegRNA.
Here we generated stem-loop PEs (sPEs) by adding stem-loop aptamers at the 3’-terminal of pegRNA
Stem-loop PEs were generated by adding stem-loop aptamers at the 3-prime terminal of pegRNA.
Here we generated stem-loop PEs (sPEs) by adding stem-loop aptamers at the 3’-terminal of pegRNA
Stem-loop PEs were generated by adding stem-loop aptamers at the 3-prime terminal of pegRNA.
Here we generated stem-loop PEs (sPEs) by adding stem-loop aptamers at the 3’-terminal of pegRNA
The modified pegRNAs were split into sgRNA and prime RNA to create split pegRNA prime editors.
We split the modified pegRNAs into sgRNA and prime RNA. The resulting split pegRNA prime editors (SnPEs)
The modified pegRNAs were split into sgRNA and prime RNA to create split pegRNA prime editors.
We split the modified pegRNAs into sgRNA and prime RNA. The resulting split pegRNA prime editors (SnPEs)
The modified pegRNAs were split into sgRNA and prime RNA to create split pegRNA prime editors.
We split the modified pegRNAs into sgRNA and prime RNA. The resulting split pegRNA prime editors (SnPEs)
The modified pegRNAs were split into sgRNA and prime RNA to create split pegRNA prime editors.
We split the modified pegRNAs into sgRNA and prime RNA. The resulting split pegRNA prime editors (SnPEs)
The modified pegRNAs were split into sgRNA and prime RNA to create split pegRNA prime editors.
We split the modified pegRNAs into sgRNA and prime RNA. The resulting split pegRNA prime editors (SnPEs)
The modified pegRNAs were split into sgRNA and prime RNA to create split pegRNA prime editors.
We split the modified pegRNAs into sgRNA and prime RNA. The resulting split pegRNA prime editors (SnPEs)
The modified pegRNAs were split into sgRNA and prime RNA to create split pegRNA prime editors.
We split the modified pegRNAs into sgRNA and prime RNA. The resulting split pegRNA prime editors (SnPEs)
Split pegRNA prime editors maintain prime editing activity and increase flexibility.
The resulting split pegRNA prime editors (SnPEs) maintain the PE activity and increase flexibility.
Split pegRNA prime editors maintain prime editing activity and increase flexibility.
The resulting split pegRNA prime editors (SnPEs) maintain the PE activity and increase flexibility.
Split pegRNA prime editors maintain prime editing activity and increase flexibility.
The resulting split pegRNA prime editors (SnPEs) maintain the PE activity and increase flexibility.
Split pegRNA prime editors maintain prime editing activity and increase flexibility.
The resulting split pegRNA prime editors (SnPEs) maintain the PE activity and increase flexibility.
Split pegRNA prime editors maintain prime editing activity and increase flexibility.
The resulting split pegRNA prime editors (SnPEs) maintain the PE activity and increase flexibility.
Split pegRNA prime editors maintain prime editing activity and increase flexibility.
The resulting split pegRNA prime editors (SnPEs) maintain the PE activity and increase flexibility.
Split pegRNA prime editors maintain prime editing activity and increase flexibility.
The resulting split pegRNA prime editors (SnPEs) maintain the PE activity and increase flexibility.
Stem-loop PEs and tethered PEs increased small insertion, deletion, or point mutation editing efficiency by 2-fold to 4-fold on average in HEK293, U2OS, and HeLa cells.
sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells.
editing efficiency fold increase 2-4 fold
Stem-loop PEs and tethered PEs increased small insertion, deletion, or point mutation editing efficiency by 2-fold to 4-fold on average in HEK293, U2OS, and HeLa cells.
sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells.
editing efficiency fold increase 2-4 fold
Stem-loop PEs and tethered PEs increased small insertion, deletion, or point mutation editing efficiency by 2-fold to 4-fold on average in HEK293, U2OS, and HeLa cells.
sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells.
editing efficiency fold increase 2-4 fold
Stem-loop PEs and tethered PEs increased small insertion, deletion, or point mutation editing efficiency by 2-fold to 4-fold on average in HEK293, U2OS, and HeLa cells.
sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells.
editing efficiency fold increase 2-4 fold
Stem-loop PEs and tethered PEs increased small insertion, deletion, or point mutation editing efficiency by 2-fold to 4-fold on average in HEK293, U2OS, and HeLa cells.
sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells.
editing efficiency fold increase 2-4 fold
Stem-loop PEs and tethered PEs increased small insertion, deletion, or point mutation editing efficiency by 2-fold to 4-fold on average in HEK293, U2OS, and HeLa cells.
sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells.
editing efficiency fold increase 2-4 fold
Stem-loop PEs and tethered PEs increased small insertion, deletion, or point mutation editing efficiency by 2-fold to 4-fold on average in HEK293, U2OS, and HeLa cells.
sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells.
editing efficiency fold increase 2-4 fold
Approval Evidence
1 source3 linked approval claimsfirst-pass slug tethered-pes
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Source:
design rationalesupports
The 3-prime extension of pegRNAs could negatively affect pegRNA stability or folding and compromise prime editing activity.
The 3’-extension of pegRNAs could negatively affect its stability or folding and comprise the PE activity.
Source:
engineering strategysupports
Stem-loop PEs can be tethered to Cas9 nickase to produce tethered PEs.
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Source:
performance improvementsupports
Stem-loop PEs and tethered PEs increased small insertion, deletion, or point mutation editing efficiency by 2-fold to 4-fold on average in HEK293, U2OS, and HeLa cells.
sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells.
Source:
Comparisons
Source-backed strengths
The reported strength of tPEs is a design rationale aimed at enhancing prime editing efficiency and flexibility. The evidence specifically supports construction of pegRNAs bearing 3-prime stem-loop aptamers that can be tethered to Cas9 nickase, but it does not provide quantitative performance data in the supplied material.
Source:
which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs)
Source:
Here we generated stem-loop PEs (sPEs) by adding stem-loop aptamers at the 3’-terminal of pegRNA
Source:
We split the modified pegRNAs into sgRNA and prime RNA. The resulting split pegRNA prime editors (SnPEs)
Source:
sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells.
Compared with intron-containing CRISPRa construct
tethered PEs and intron-containing CRISPRa construct address a similar problem space because they share editing, recombination.
Shared frame: same top-level item type; shared target processes: editing, recombination
Compared with split pegRNA prime editors
tethered PEs and split pegRNA prime editors address a similar problem space because they share editing, recombination.
Shared frame: same top-level item type; shared target processes: editing, recombination; shared mechanisms: prime editing
Compared with stem-loop PEs
tethered PEs and stem-loop PEs address a similar problem space because they share editing, recombination.
Shared frame: same top-level item type; shared target processes: editing, recombination; shared mechanisms: rna-protein tethering
Ranked Citations
- 1.