Perovskite light-emitting diodes (LEDs) have attracted broad attention due to their rapidly increasing external quantum efficiencies (EQEs) [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15] . However, most high EQEs of perovskite LEDs are reported at low current densities (< 1 mA cm -2 ) and low brightness. Decrease in efficiency and rapid degradation at high brightness inhibit their practical applications. Here,
CRISPR/Cas9 genome-editing tools have tremendously boosted our capability of manipulating the eukaryotic genomes in biomedical research and innovative biotechnologies. However, the current approaches that allow precise integration of gene-sized large DNA fragments generally suffer from low efficiency and high cost. Herein, we developed a versatile and efficient approach, termed LOCK (
L
ong dsDNA with 3′-
O
verhangs mediated
C
RISPR
K
nock-in), by utilizing specially designed 3′-overhang double-stranded DNA (odsDNA) donors harboring 50-nt homology arm. The length of the 3′-overhangs of odsDNA is specified by the five consecutive phosphorothioate modifications. Compared with existing methods, LOCK allows highly efficient targeted insertion of kilobase-sized DNA fragments into the mammalian genomes with low cost and low off-target effects, yielding >fivefold higher knock-in frequencies than conventional homologous recombination-based approaches. This newly designed LOCK approach based on homology-directed repair is a powerful tool suitable for gene-sized fragment integration that is urgently needed for genetic engineering, gene therapies, and synthetic biology.
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