Cell culture-adaptive mutations in hepatitis C virus promote viral production by enhancing viral replication and release

Table 1 Sequence of primers used for adaptive mutation plasmid construction

Primer	Sequence (5’-3’)
1340-F	CTGGCGTACGTGATGCG
m2310-R	TGTCCCTGTCCTCCAAGCCGCAGCGAT
m2310-F	ATCGCTGCGGCTTGGAGGACAGGGACA
3500-R	GCCCCGTCATACTCACCAC
m2681-R	TGATGTACCAAGCTGCCACGAAGAAG
m2681-F	CTTCTTCGTGGCAGCTTGGTACATCA
5249-F	AATGAGGTCACCCTCACACA
m6132-R	ACGTGGCTTCCTCTGGAAGCAAAGGCA
m6132-F	TGCCTTTGCTTCCAGAGGAAGCCACGT
7791-R	GATGTTGTACAGTACACCTTG
5249-F	AATGAGGTCACCCTCACACA
m7160-R	AGCATGCGCTCCGATGGTATTGAG
m7160-F	CTCAATACCATCGGAGCGCATGCT
m7359-R	TTCTGATGTGGTGCTCTCGCTCAG
m7359-F	CTGAGCGAGAGCACCACATCAGAA
m7658-R	GCACAGGGTGGTATCGTCCTCCT
m7658-F	AGGAGGACGATACCACCCTGTGC
7966-R	CTTGGATCTTGCAGAAT

Table 2 Primer combinations used in adaptive mutation plasmid construction

Fragment	Template	Primers
		Sense	Anti-sense
mE2-1	JFH1	1340-F	m2310-R
mE2-2		m2310-F	3500-R
mp7-1		1340-F	m2681-R
mp7-2		m2681-F	3500-R
mNS4B-1		5249-F	m6132-R
mNS4B-2		m6132-F	7791-R
mNS5A-1		5249-F	m7160-R
mNS5A-2		m7160-F	m7359-R
mNS5A-3		m7359-F	m7658-R
mNS5A-4		m7658-F	7966-R
mNS4B/NS5A-1	JFH1-mNS5A	5249-F	m6132-R
mNS4B/NS5A-2		m6132-F	7791-R
mE2/p7 -1	JFH1-mp7/NS5A	1350-F	m2310-R
mE2/p7 -2		m2310-F	3500-R

Table 3 Primers and templates for overlap PCR

Fragment	Template	Primer
		Upstream	Downstream
mE2	mE2-1 + mE2-2	1340-F	3500-R
mp7	mp7-1 + mp7-2	1340-F	3500-R
mNS4B	mNS4B-1 + mNS4B-2	5249 F	7791-R
mNS5A-3/4	mNS5A-3 + mNS5A-4	m7359-F	7966-R
mNS5A-2/3/4	mNS5A-2 + mNS5A-3/4	m7160-F	7966-R
mNS5A	mNS5A-1 + mNS5A-2/3/4	5249-F	7966-R
mNS4B/NS5A	mNS4B/NS5A-1 + mNS4B/NS5A-2	5249 F	7791-R
mE2/p7	mE2/p7 -1 + mE2/p7-2	1340-F	3500-R