TABLE 1.
Plasmid | Promoter | RBSb | 5′ additionc | No. of codons after landing sited | U of activity withe:
|
Avg bypassing efficiency ± SD (%)f | |
---|---|---|---|---|---|---|---|
Test fusion | Δgap fusion | ||||||
BX3* | tac | UAGAGGGUAUUAUG | AUGAAAAGCUUA(AUG) | 2 | 6,107 ± 490 | 24,270 ± 2,540 | 24.7 ± 1.4 |
pSK31 | tac | Same as for BX3* | GST | 7 | 4,432 ± 85 | 31,648 ± 1,956 | 14.0 ± 0.8 |
pPG33 | tac | Same as for BX3* | T7 gene 10 | 7 | 5,042 ± 120 | 42,156 ± 1,895 | 12.0 ± 0.5 |
pPA31 | tac | Same as for BX3* | ProtA-CAT | 7 | 11,484 ± 590 | 49,852 ± 1,890 | 23.0 ± 1.4 |
NH11 | λ pLpR | bla signal peptide | 24 | 81 ± 6 | 216 ± 12 | 37 ± 3 | |
GLZ16 | Consensus | UCAGGAAGCUUAUG | 2 | 140 ± 3 | 2,273 ± 27 | 6.2 ± 0.1 | |
GLZ22 | Consensus | Same as for GLZ16 | 24 | 99 ± 5 | 960 ± 28 | 10.3 ± 0.3 | |
GLZ30 | tac | Same as for GLZ16 | 24 | 7,010 ± 1,400 | 18,580 ± 1,170 | 37 ± 4 | |
GLZ32 | Consensus | UCCGGAAGCUUAUG | 24 | 69 ± 2 | 706 ± 20 | 9.8 ± 0.3 | |
GLZ34 | Consensus | UGAGGAAGCUUAUG | 24 | 209 ± 16 | 1,320 ± 60 | 15.9 ± 0.6 | |
GLZ40 | Consensus | AUGAGGUGUAUAAUG | 24 | 207 ± 24 | 1,370 ± 35 | 15.2 ± 1.4 | |
GLZ49 | Consensus | UAGGAGGAAGCUUAUG | 24 | 131 ± 5 | 919 ± 45 | 14.3 ± 0.2 | |
pGG34 | tac | CAGGAAACAGAAGCUUAUG | 2 | 19,220 ± 3,540 | 72,450 ± 6,232 | 26 ± 5 | |
pGG36 | lacUV5 | Same as for pGG34 | 2 | 4,250 ± 820 | 23,620 ± 2,750 | 18 ± 4 | |
pGG45 | tac | Same as for pGG34 | 7 | 10,940 ± 950 | 52,080 ± 3,820 | 21 ± 2 | |
pGG47 | tac | Same as for pGG34 | 24 | 14,200 ± 1,620 | 50,640 ± 4,520 | 28 ± 4 |
All reporter fusions were made to lacZ except that with NH11, which was made to phoA.
The underlined region is the SD region of the ribosome-binding sequence (RBS); the boldface region is the initiation codon in each construct.
In the sequence, the initiation codon is in boldface type, and the beginning of the gene 60 coding sequence is in parentheses. T7 gene 10, 5′ fragment of T7 gene 10; ProtA, immunoglobin-binding domain from protein A. BX3* carries two transversions at nt 87 and 89 that create an internal BglII site and that do not affect bypassing efficiency.
Number of gene 60 codons between the landing site and reporter fusion.
Activities are expressed in β-Gal units, except for that for NH11, which is expressed in phosphatase units. Values are averages of multiple independent measurements. Δgap, gap deletion.
Values are average efficiency estimates from several independent experiments.