Peptide/Cas9 nanostructures for ribonucleoprotein cell membrane transport and gene edition (2017)
PT24 (oleic aldehyde hydrazone amphiphile from peptide P)
Sequence: Arg/Leu-rich cationic peptide scaffold with two hydrazide (Hy) residues (exact sequence not in main text; schematic shows RRLKRRLLRRLKRL)
gRNA (CRISPR guide RNA) in Cas9 RNP
| Experiment Id | EXP001805 |
|---|---|
| Paper | Peptide/Cas9 nanostructures for ribonucleoprotein cell membrane transport and gene edition |
| Peptide | PT24 (oleic aldehyde hydrazone amphiphile from peptide P) |
| Delivery Success Class | no |
| In Vivo Flag | no |
| Uptake Confirmed | yes |
| Label Confidence | medium |
| In Vitro Functional Effect | yes |
| Endosomal Escape Evidence | yes |
| Peptide Concentration | PT24 10 mM (optimized); also tested 0.5–12.5 µM for gel retardation; uptake studies at 5 µM TmPT24 |
| Rna Concentration | Cas9 RNP 40–160 nM (dose-response); typical 80 nM or 160 nM; peptide:RNP ratio 62.5:1 (mol/mol) used in T7E1 assays |
| Mixing Ratio | RNP:PT24 = 1:62.5 (mol/mol) (editing assays) |
| Formulation Format | Non-covalent peptide amphiphile/Cas9 ribonucleoprotein nanoparticles |
| Formulation Components | PT24 amphiphilic CPP + SpCas9 protein complexed with gRNA (RNP); TAMRA-labelled PT24 (TmPT24) for uptake; DexPT24 for GR translocation assay |
| Size Nm | 270.00 |
| Zeta Mv | 4.20 |
| Model Scope | in_vitro |
| Model Type | in vitro |
| Cell Lines Or Primary Cells | HeLa; A549; DF1 (chicken fibroblast); DF1-EGFP |
| Animal Model | |
| Administration Route | |
| Output Type | Genome editing (T7E1 indel % at 48 h; multiple cell lines) |
| Output Value | HeLa HPRT1 T7E1 up to ~38% (160 nM RNP) with PT24 (similar to Lipofectamine); ~26% in 5% serum; A549 HPRT1 ~21% indels; DF1-EGFP ~39% EGFP indels |
| Output Units | |
| Output Notes | Uptake mainly macropinocytosis (EIPA-sensitive); lysosomal disruption (LysoTracker decrease) and GR–GFP nuclear translocation with DexPT24 confirm cytosolic access/endosomal escape. |
| Toxicity Notes | Lower toxicity vs Lipofectamine 2000 at comparable editing (MTT). |
| Curation Notes |