Amplification editing empowers in situ large-scale DNA duplication

Repetitive sequences in genomes play crucial biological roles involved in maintaining chromosome structure or the development of diseases. Modeling naturally occurring large-scale repetitive sequences will facilitate the exploration of their functions and elucidate their underlying mechanisms. However, constructing programmable large-scale repetitive sequences on chromosomes remains a challenge. The Yin laboratory’s latest study published in Cell reported a significant breakthrough in the generation of large-scale repetitive sequences on chromosomes, with the method demonstrating feasibility across various human- and mouse-derived cell lines, including diploid, haploid, primary, and embryonic stem cells (ESC) (Figure 1).1 The researchers ingeniously altered the recognition region of nCas9 from a PAM-in orientation (where the PAM sequences are within the target region) to a PAM-out orientation (where the PAM sequences are at the end of the target region), resulting in in situ tandem duplication of chromosomal segments. Notably, they achieved 100 Mb replication at the chromosomal scale for the first time. Furthermore, amplification editing (AE) can generate multiple rounds of replication, producing tandem repeat sequences that range from 20 bp to 8 kb with fewer insertion-deletion mutations (indels).