I am relatively new to cloning. I want to design a neomycin cassette (dsODN) with specific 40-60bp overhangs (one overhang on either stranded). I want to transfect my cassette in with the CRISPR nickase that cleaves single stranded DNA about 40nt apart (exacly like Figure 5a in Cell 154, 1380–1389, September 12, 2013, except with neomycin)
I have no idea how I would go about doing an overhang PCR reaction for homologous arms. Could some help me or provide a protocol?