text: Scar sequences are common in multiple segment DNA assembly. In the multisegment assembly method Gateway, segments are added into the donor with additional ATT sequences, which overlap in those added segments, and this results in the segments separated by the ATT sequences. In BioBrick assembly, an eight-nucleotide scar sequence, which codes for a tyrosine and a stop codon, is left between every segment added into the plasmid.
  Golden Gate assembly uses Type IIS restriction enzymes cutting outside their recognition sequences. Also, the same Type IIS restriction enzyme can generate copious different overhangs on the inserts and the vector; for instance, BsaI creates 256 four-basepair overhangs. If the overhangs are carefully designed, the segments are ligated without scar sequences between them, and the final construct can be quasi-scarless, where the restriction enzyme sites remain on both sides of the insert. As additional segments can be inserted into the vectors without scars within an open reading frame, Golden Gate is widely used in protein engineering.