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. 2006 Apr;74(4):2093–2101. doi: 10.1128/IAI.74.4.2093-2101.2006

TABLE 2.

VacA expression plasmids generated in this study

Expression plasmida Forward primerb Reverse primerb Restriction enzymesc Encoded VacA proteind
pMM652 AND3817 AND6003 SpeI/PstI Δ(6-27)1-821-His
pVT562 AND3817 OP9133 SpeI/SalI Δ(6-27)1-800
pVT561 AND3817 OP9135 SpeI/SalI Δ(6-27)1-780
pVT560 AND3817 BAR1559 SpeI/SalI Δ(6-27)1-700
pVT559 AND3817 BAR1558 SpeI/SalI Δ(6-27)1-550
pVT315 AND3817 AND6001 SpeI/PstI Δ6-27p48-His
pVT148 AND3817 OP6228 SpeI/PstI Δ6-27p33-His
pVT587 OPE1144 OPE1145 NAe Δ(6-27/334-360)p48-His
pVT604 OPE1216 OPE1217 NAe Δ(6-27/334-341)p48-His
pVT606 OPE1218 OPE1219 NAe Δ(6-27/342-350)p48-His
pVT608 OPE1220 OPE1221 NAe Δ(6-27/351-360)p48-His
pVT597 OPE1216 OPE1217 NAe Δ334-341p55-His
pVT598 OPE1218 OPE1219 NAe Δ342-350p55-His
pVT600 OPE1220 OPE1221 NAe Δ351-360p55-His
a

Each of these plasmids was derived from pET41b.

b

Oligonucleotides used to PCR amplify the different vacA sequences. Oligonucleotide sequences are listed in Table 1.

c

Restriction sites used to clone the PCR products into pET41b.

d

The VacA amino acid numbering system used in this table is based on designating the first amino acid (alanine) of the mature secreted VacA toxin of strain 60190 as amino acid 1. “Δ” indicates amino acids that are deleted from the mutant VacA fragments. “His” indicates the presence of a six-His tag. All of these VacA proteins that contain a Δ6-27 deletion also contain an alanine-to-methionine substitution at amino acid 1 (A1M) (26).

e

NA, not applicable. These plasmids containing in-frame internal deletions were constructed by inverse PCR as described in Materials and Methods.