Name
|
Type
|
Primer sequences
|
Primer sequences
|
Restriction site
|
---|
F-Ras1a
|
Forward
|
rlmlrab
|
5′-GCCCATATGAGCTCAACTGGTCAGCATG-3′
|
NdeI
|
F-Ras3a
|
Forward
|
rlmlrabC
|
5′GCCCATATGATCATCGCCGATGTGTCG-3′
|
NdeI
|
R-Ras3b
|
Reverse
|
rlmlrab/rlmlrabC
|
5′-GCCGCGGCCGCCCAGTACGCAGCAGTTGC-3′
|
NotI
|
- Reverse and forward primers containing at their 5′ end a restriction site (underlined) were used to amplify the full length LmlRAB ORF and its carboxyl part LmlRABC. The amplified double-stranded DNA were first digested with NdeI and NotI and then inserted into the corresponding cloning sites in the pET-22b + vector (Novagen)