Reagent	Amount
5× Phusion HF Buffer	10 μL
10 mM dNTPs	1 μL
no-homology HYG DNA forward primer (10 μM)	2.5 μL
no-homology HYG DNA reverse primer (10 μM)	2.5 μL
Phusion DNA Polymerase	0.5 μL
pFGL821 (∼5 ng/μL)	1 μL
Molecular water	32.5 μL
Total	50 μL