Efficient Protein Expression System for Expressing Bt Insecticidal Protein

TECHNICAL FIELD

The present invention relates to the field of protein (in particular, Bt protein) expression. The present invention provides an efficient protein expression system for expressing a Bt insecticidal protein. The protein expression system uses Bla (beta-lactamase) as a fusion tag, so that Bt genes unable to be expressed as soluble proteins are expressed efficiently as soluble proteins.

BACKGROUND ART

In vitro expression technology of a protein is an important part of genetic engineering technology. Compared with other protein expression systems, Escherichia coli is an efficient and economical way, and the expression level of a recombinant protein can reach 50% of the total protein content of Escherichia coli. The recombinant protein with a normal biochemical activity is usually in a soluble form. Therefore, in order to obtain active proteins, soluble expression pathways are usually used.

A Bt protein is a parasporal crystal protein of Bacillus thuringiensis formed in bacteria during a sporulation period. The Bt protein is a widely used biopesticide, which is safe and has a high-efficiency and broad-spectrum insecticidal activity, and is harmless to humans, animals and non-target pests. Exploring new Bt proteins has very high commercial application value.

Expresso® Solubility & Expression Screening System from Lucigen is a protein expression tag screening kit containing 7 fusion tags. Gene fragments can be conveniently connected to 8 different vectors through homologous recombination reactions. Through the rapid screening of 7 different fusion tags, the most suitable tag for expressing the target protein is obtained (see Table 1 below).

TABLE 1

#
Fusion tag
AA length
kDa*
pI
Description

1
6xHis-AFV
113
13.5
5.0
Hypothetical protein from

Acidianus filovirus 1

2
6xHis-SlyD
210
22.7
5.2
FKBP-type peptidylprolyl

cis-trans isomerase

3
6xHis-Tsf
297
32.2
5.7

E. coli elongation factor

4
6xHis-SUMO
115
13.3
5.2
Small ubiquitin-like

modifier

5
6xHis-Bla
381
41.3
4.4
Beta-lactamase

6
6xHis-MBP
382
42.1
5.5
Maltose-binding protein

7
6xHis-GST
233
27.4
6.6
Glutathione S-transferase

8
6xHis control
14
1.8
7.0
Affinity tag

The newly explored Bt genes need to be expressed as soluble proteins to test the insecticidal activity thereof. However, relying on the current Escherichia coli expression technology, only about 50% to 60% of Bt genes can be successfully expressed. Therefore, the successful expression of Bt genes as soluble proteins has important research and commercial value.

SUMMARY OF THE INVENTION

The present invention establishes an efficient protein expression system. Using the existing Escherichia coli expression technology, about 50% to 60% of Bt genes can be successfully expressed as soluble proteins. Using the method of the present invention, the remaining 40% to 50% of Bt genes unable to be expressed as soluble proteins can achieve 80% successful expression (expressed as soluble proteins), and the total success rate of Bt gene expression can reach at least 80%.

In the present invention, 16 Bt genes unable to be expressed as soluble proteins were initially used as test samples and screened for fusion tags using 8 vectors in Expresso® Solubility & Expression Screening System from Lucigen. The results showed that the pSol-Bla vector could achieve 100% expression for 16 test samples. For the convenience of cloning, we modified the pSol-Bla vector from Lucigen and used the modified vector to express an additional 237 Bt genes, which are unable to be expressed as soluble proteins, in Escherichia coli; as a result, 193 genes were successfully expressed as soluble proteins, and the expression success rate was 81.4%. Therefore, for a total of 253 (16+237) Bt genes unable to be expressed as soluble proteins before, 209 (16+193) genes were successfully expressed as soluble proteins, and the expression success rate was 82.6%.

In one aspect, the present invention relates to the use of Bla, i.e., beta-lactamase, as a fusion tag for expressing a Bt protein.

In another aspect, the present invention relates to the use of a modified pSol-Bla vector as shown in FIG. 1 for expressing a Bt protein.

Preferably, in the above-mentioned uses, the Bt protein is expressed in Escherichia coli. More preferably, the Escherichia coli is Escherichia coli 10G or BL21 (DE3). Preferably, in the above-mentioned uses, the Bt protein is a Bt protein unable to be expressed.

In another aspect, the present invention relates to a fusion protein comprising a Bt protein and a beta-lactamase as a fusion tag. Preferably, the Bt protein is a Bt protein unable to be expressed.

In another aspect, the present invention relates to a nucleic acid molecule encoding the above-mentioned fusion protein.

In another aspect, the present invention relates to a vector comprising the above-mentioned nucleic acid molecule. Preferably, the vector is a pSol-Bla vector into which a Bt gene to be expressed is inserted, or a modified pSol-Bla vector as shown in FIG. 1 into which a Bt gene to be expressed is inserted.

In another aspect, the present invention relates to a cell comprising the above-mentioned vector. Preferably, the cell is an Escherichia coli cell. More preferably, the Escherichia coli cell is an Escherichia coli 10G or BL21 (DE3) cell.

In another aspect, the present invention relates to a method for expressing a Bt protein, comprising: (1) culturing the above-mentioned cell under conditions suitable for expressing the desired protein; and (2) optionally, cleaving the Bla tag to obtain a Bt protein without the tag.

In another aspect, the present invention relates to a modified pSol-Bla vector as shown in FIG. 1.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a map showing the modified pSol-Bla vector of the present invention.

DETAILED DESCRIPTION OF EMBODIMENTS

The embodiments of the present invention will be described in detail below in combination with examples, but a person skilled in the art would understand that the following examples serve only to illustrate the present invention and are not intended to limit the scope of the present invention. If the specific conditions are not indicated in the examples, the conventional conditions or the conditions suggested by the manufacturer shall be followed. Any reagents or instruments used, unless the manufacture stated, are conventional products that can be obtained by market purchase.

Example 1. Screening of Fusion Tags for 16 Bt Genes Unable to be Expressed as Soluble Proteins to Achieve Soluble Expression

We selected 16 Bt genes unable to be expressed as soluble proteins through conventional methods (for example, the expression of pET series vectors at different temperatures and IPTG induced concentrations, the use of fusion tags such as SUMO and GST, the use of different types of competent cells, cell-free culture, expression with Bt cells, etc.). Their numbers, molecular weights, and expression issues are shown in Table 2 below:

TABLE 2

Protein

Molecular
Expression

number
Gene family
SEQ ID NO
weight
issue

BT-0025
Cry1If
1
80
KD
Insoluble

BT-0027

2
79
KD
No expression

BT-0275
Cry22Aa
3
96
KD
No expression

BT-0362

4
73
KD
No expression

BT-0425
Cry36Aa
5
56
KD
No expression

BT-0435
Cry42Aa
6
75
KD
No expression

BT-0445
Vip1Aa
7
138
KD
Insoluble

BT-0452
Cry19Ca
8
59
KD
No expression

BT-0560
Cry20Aa
9
26
KD
No expression

BT-0587
Cry49Ab
10
60
KD
No expression

BT-0669
Cry2Ae
11
30
KD
Insoluble

BT-0685
Cry49Ab
12
51
KD
Insoluble

BT-0827
Cry48Ab2
13
86
KD
No expression

BT-0830
Cry48Ab2
14
87
KD
No expression

BT-0831
Cry54Ba1
15
80
KD
No expression

HO4
Cry1Ab.1Ca
16
130
KD
Insoluble

Using Expresso® Solubility and Expression Screening System from Lucigen, according to the instructions thereof, the primer sequences at both ends were designed to synthesize PCR primers of these 16 genes, respectively. The target fragments of 16 genes with homologous recombination ends were obtained by PCR amplification methods. Then, after mixing the PCR products with 8 linear vectors in the kit, the chemically competent cells were transformed with plasmids. After the positive clones were identified from the obtained monoclonal plasmids by PCR, the positive clones were sent to a company for sequencing and identification. After analyzing the sequencing results, we obtained the plasmids of these 16 genes on 8 vectors, a total of 128 plasmids.

30 μl of Escherichia coli 10G or BL21 (DE3) glycerol bacteria with these 16 genes were cultured in 3 ml of LB medium at 37° C. to an OD600 of approximately 0.5, and then 20% rhamnose was added to induce culture overnight at 18° C. The bacteria were collected and subjected to disruption by sonication, and whole bacteria and supernatant were collected respectively and boiled for 10 minutes before the expression was analyzed by SDS-PAGE. The expression of these 128 plasmids in Escherichia coli 10G and BL21 (DE3) cells was shown in Table 3 below (the first 6 genes in Table 3 were purified after successful expression and the Bla tag was cleaved to obtain a pure soluble target protein).

TABLE 3

Issue when

expressing

through

conventional
Valid tag

Protein
methods
10 G
BL21 (DE3)

BT-0025 (80 KD)
Insoluble
6 tags (Bla, GST,
6 tags (Bla, GST,

His, MBP, SUMO, Tsf)
His, MBP, SUMO, Tsf)

BT-0827 (86 KD)
No
4 tags (AFV
Not tested

expression
Bla, GST, Tsf)

BT-0830 (87 KD)
No
5 tags (Bla, GST,
Not tested

expression
MBP, Tsf, SlyD)

BT-0831 (80 KD)
No
6 tags (AFV, Bla,
Not tested

expression
GST, MBP, SlyD, Tsf)

HO4 (130 KD)
Insoluble
4 tags (AFV,
Not tested

Bla, MBP, Tsf)

BT-0027 (79 KD)
No
3 tags (Bla,
Not tested

expression
MBP, SlyD)

BT-0685 (51 KD)
Insoluble
All 8 tags
5 tags (AFV, Bla,

GST, MBP, His)

BT-0669 (30 KD)
Insoluble
7 tags (All,
7 tags (All

except for GST)
except for GST)

BT-0587 (60 KD)
No
6 tags (AFV, Bla,
All 8 tags

expression
GST, His, MBP, SUMO)

BT-0560 (26 KD)
No
3 tags (Bla, MBT, Tsf)
4 tags (AFV,

expression

GST, MBP, SUMO)

BT-0362 (73 KD)
No
4 tags (Bla, MBP,
Not tested

expression
SlyD, Tsf)

BT-0425 (56 KD)
No
5 tags (AFV, Bla,
Not tested

expression
MBP, SUMO, Tsf)

BT-0452 (59 KD)
No
5 tags (AFV, Bla,
Not tested

expression
GST, MBP, SlyD)

BT-0435 (75 KD)
No
2 tags (AFV, Bla)
Not tested

expression

BT-0445 (138 KD)
Insoluble
4 tags (AFV,
4 tags (AFV, Bla,

Bla, MBP, Tsf)
MBP, Tsf)

BT-0275 (96 KD)
No
2 tags (AFV, Bla)
Not tested

expression

The results showed that most of the 8 vectors with different tags in the kit could express individual genes in these 16 Bt genes, and only the pSol-Bla vector was effective for all 16 genes (that is, the genes could be expressed as soluble proteins), implying that the Bla tag specifically improved the effect of the Bt gene expression.

Example 2. Modification of pSol-Bla Vector from Lucigen and Use of the Modified pSol-Bla Vector on 237 Bt Genes Unable to be Expressed as Soluble Proteins

For the convenience of cloning, we made a few modifications to the pSol-Bla vector from Lucigen, wherein the stop codon TAA at positions 1326-1328 in the original vector was removed, and the sequence “ggatccggctcaggctcaggctcaggctcaggctcaggctcactcgag” was added at position 1326 of the original vector, the sequence comprising one BamHI enzyme digestion site and one XhoI enzyme digestion site. The modified pSol-Bla vector was as shown in FIG. 1. We selected 237 Bt genes unable to be expressed as soluble proteins (using a pET28a vector, the soluble form of protein cannot be obtained in the supernatant of the cell lysate under conditions of induction overnight with 0.2-0.5 mM IPTG at 18° C.), and cloned the genes into the modified pSol-Bla vector through enzyme digestion and ligation. After transforming the vector into Escherichia coli 10G cells, we used the same method to express these 237 plasmids: 30 μl of glycerol bacteria were added to 3 ml of LB medium and cultured at 37° C. to an OD600 of approximately 0.5, and then 20% rhamnose was added to induce the culture overnight at 18° C. The bacteria were collected and subjected to disruption by sonication, and whole bacteria and supernatant were collected respectively and boiled for 10 minutes before the expression was analyzed by SDS-PAGE.

The results showed that 193 out of the 237 Bt genes unable to be expressed as soluble proteins obtained soluble fusion proteins in this way, and the success rate was up to 81.4%. This result showed that the vector fused with Bla could specifically increase the success rate of the successful expression of the Bt gene.

The information regarding the 237 Bt genes unable to be expressed as soluble proteins is as follows:

TABLE 4

Molecular

Protein

weight
Whether expressed

number
Gene family
SEQ ID NO
(KD)
in this experiment

BT-0269
Cry41Ab
17
93
Yes

BT-0290
Cry19Ca
18
136
No

BT-0397
Cry59Aa
19
77
Yes

BT-0398
Cry4Ca
20
75
Yes

BT-0411
Vip2Ad
21
22.5
Yes

BT-0426
Cry59Aa
22
73
Yes

BT-0434
Cry21Ba
23
120
Yes

BT-0439
Cry36Aa
24
57
Yes

BT-0441
Cry68Aa
25
82
Yes

BT-0455
Cry41Ba
26
94
Yes

BT-0456
Cry22Ab
27
79
Yes

BT-0464
Cry44Aa
28
59
Yes

BT-0474
Cry41Ba
29
92
Yes

BT-0481
Cry24Ca
30
45
Yes

BT-0482
Cry4Cc
31
49
Yes

BT-0483
Cry4Ca
32
131
Yes

BT-0526
Cry2Aa
33
39
Yes

BT-0532
Vip2Af
34
50
Yes

BT-0552
Cry15Aa
35
34
Yes

BT-0555
Cry52Aa
36
77
Yes

BT-0557
Cry53Ab
37
74
No

BT-0567
Cry70Bb
38
106
Yes

BT-0579
Cry40Da
39
74
Yes

BT-0581
Cry40Da
40
72
Yes

BT-0585
Cry36Aa
41
52
Yes

BT-0586
Cry49Aa
42
56
Yes

BT-0591
Cry4Cc
43
78
No

BT-0596
Cry4Ca
44
61
Yes

BT-0597
Cry69Aa
45
135
No

BT-0599
Cry54Aa
46
72
Yes

BT-0600
Cry56Aa
47
70
Yes

BT-0603
Cry53Ab
48
76
Yes

BT-0609
Cry4-like
49
26
Yes

BT-0610
Cry68Aa
50
89
Yes

BT-0612
Cry15Aa
51
40
No

BT-0613
Cry51Aa
52
37
Yes

BT-0624
Cry22Ab
53
64
Yes

BT-0626
Cry49Aa
54
23
Yes

BT-0663
Cry68Aa
55
82
Yes

BT-0666
Cry68Aa
56
90
Yes

BT-0667
Cry68Aa
57
91
Yes

BT-0671
Cry14Ab
58
90
Yes

BT-0681
Cry42Aa
59
88
Yes

BT-0683
Cry19Ca
60
76
Yes

BT-0713
Cry41Aa1
61
56
Yes

BT-0735
Cry4Cb1
62
79
Yes

BT-0744
Cry60Aa
63
35
Yes

BT-0760
Cry64Aa1
64
37
No

BT-0762
Cry64Aa1
65
36
No

BT-0764
Cry15Aa1
66
34
Yes

BT-0766
Cry54Aa2
67
78
Yes

BT-0767
Cry20Ba1
68
83
Yes

BT-0768
Cry51Aa1
69
35
No

BT-0770
Cry51Aa1
70
40
Yes

BT-0772
Cry70Bb1
71
110
No

BT-0776
Cry32Mb1
72
138
No

BT-0777
Cry32Ra1
73
126
Yes

BT-0778
Cry32Cb1
74
96
Yes

BT-0781
cry69Aa1
75
81
Yes

BT-0783
Cry40Da1
76
74
Yes

BT-0784
Cry4Ca1
77
65
No

BT-0785
Cry55Aa1
78
35
Yes

BT-0787
Cry55Aa1
79
35
No

BT-0788
Cry60Aa1
80
36
Yes

BT-0795
Cry41Aa1
81
66
Yes

BT-0878
Cry4Ca1

66
Yes

BT-0879
Cry40Da1

74
Yes

BT-0895
Cry49Ab1

43
Yes

BT-0903
Vip2Ba2

44
Yes

BT-0905
Vip2Ac1

58
Yes

BT-0911
Cry14Aa1
82
91
Yes

BT-0912
Cry32Ta1
83
147
Yes

BT-0914
Cry73Aa1
84
89
Yes

BT-0915
Cry32Sa1
85
139
Yes

BT-0916
Cry41Ba2
86
86
Yes

BT-0920
Cry21Ca1
87
146
Yes

BT-0922
Cry4Aa4
88
137
Yes

BT-0927
Cry8Ga3
89
37
Yes

BT-0934
Cyt2Ca1
90
25
Yes

BT-0936
Cry49Ab1
91
31
Yes

BT-0938
Cry49Ab1
92
42
Yes

BT-0939
Cry13Aa1
93
68
Yes

BT-0941
Cry32Ra1
94
155
No

BT-0942
Cry60Aa3
95
36
Yes

BT-0943
Cry8La1
96
115
Yes

BT-0944
Cry30Db1
97
78
Yes

BT-0946
Cry51Aa2
98
32
Yes

BT-0948
Cry32Da1
99
138
Yes

BT-0949
Cry8Ca1
100
132
Yes

BT-0950
Cry60Ba3
101
36
Yes

BT-0951
Cry32Ra1
102
147
Yes

BT-0952
Cry73Aa1
103
59
Yes

BT-0953
Cry73Aa1
104
57
Yes

BT-0954
Cry45Aa
105
31
Yes

BT-0955
Cry32Qa1
106
150
Yes

BT-0956
Cry20Aa1
107
81
Yes

BT-0957
Cry54Aa2
108
77
Yes

BT-0958
Cry60Aa3
109
36
Yes

BT-0961
Cry41Ba2
110
63
No

BT-0962
Cry19Ca1
111
86
Yes

BT-0964
Cry28Aa2
112
125
Yes

BT-0965
Cry60Ba3
113
35
Yes

BT-0968
Cry43Ca1
114
145
Yes

BT-0970
Cry4Ca1
115
139
Yes

BT-0976
Cry8Ib1
116
137
No

BT-0977
Cry22Ba1
117
53
Yes

BT-0979
Cry42Aa1
118
57
Yes

BT-0980
Cry30Aa1
119
78
Yes

BT-0981
Cry4Ca1
120
136
Yes

BT-0983
Cry54Aa2
121
80
Yes

BT-0984
Cry4Cb2
122
129
Yes

BT-0985
Cyt2Ca1
123
29
Yes

BT-0987
Cry36Aa1
124
54
Yes

BT-0989
Cry4Ca1
125
64
Yes

BT-0991
Cry40Da1
126
76
Yes

BT-0993
Cry70Bb1
127
113
Yes

BT-0997
Cry8Ia1
128
139
No

BT-0998
Cry45Aa
129
29
Yes

BT-0999
Cry45Aa
130
30
Yes

BT-1000
Cry32Ra1
131
151
Yes

BT-1002
Cry11Bb1
132
33
Yes

BT-1003
Cry56Aa2
133
78
Yes

BT-1004
Cry53Ab1
134
76
Yes

BT-1006
Cry60Ba3
135
35
Yes

BT-1008
Cry60Ba3
136
35
Yes

BT-1009
Cry10Aa4
137
82
No

BT-1010
Cry9Ga1
138
84
Yes

BT-1013
Cry32Ca1
139
145
Yes

BT-1015
Cry32Qa1
140
143
No

BT-1018
Vip1Ba2
141
88
Yes

BT-1019
Cry60Ba3
142
32
Yes

BT-1020
Cry2Ah1
143
57
Yes

BT-1021
Cry70Bb1
144
109
Yes

BT-1022
Cry30Ga2
145
81
No

BT-1023
Cry39Aa1
146
78
Yes

BT-1026
Cry56Aa2
147
73
Yes

BT-1027
Cry72Aa1
148
79
Yes

BT-1028
Cry30Fa1
149
79
Yes

BT-1029
Cyt2Aa4

29
No

BT-1031
Cry40Da1
150
86
Yes

BT-1032
Cry4Ca1
151
64
Yes

BT-1033
Cry73Aa1
152
64
Yes

BT-1034
Vip2Ae2
153
84
Yes

BT-1036
Cry32Qa1
154
143
Yes

BT-1037
Cry54Ba1
155
34
Yes

BT-1042
Cry4Ba5
156
134
Yes

BT-1045
Cry67Aa2
157
125
Yes

BT-1048
Cry32Cb1
158
144
Yes

BT-1050
Vip3Ad1
159
104
Yes

BT-1051
Cry73Aa1
160
54
Yes

BT-1054
Cry2Aa3
161
36
Yes

BT-1055
Cry73Aa1
162
58
Yes

BT-1057
Cry15Aa1
163
38
Yes

BT-1060
Cry4Ba5
164
140
No

BT-1062
Cry8Aa1
165
77
Yes

BT-1064
Cry8Ga3
166
129
Yes

BT-1065
Cry29Aa1
167
78
Yes

BT-1068
Cry51Aa2
168
32
Yes

BT-1069
Cry70Bb1
169
93
Yes

BT-1070
Cry4Aa4
170
134
No

BT-1073
Cry32La1
171
137
No

BT-1076
Cry60Aa3
172
40
No

BT-1078
Cry22Aa3
173
33
Yes

BT-1084
Cry33Aa1
174
30
Yes

BT-1085
Cry32La1
175
128
Yes

BT-1087
Cry32La1
176
75
Yes

BT-1088
Cry73Aa1
177
91
Yes

BT-1090
Cyt2Aa4
178
31
Yes

BT-1091
Cry32Ra1
179
151
No

BT-1092
Cry51Aa1
180
35
Yes

BT-1093
Vip1Aa2
181
88
Yes

BT-1094
Cry73Aa1
182
94
Yes

BT-1095
Cry32Ra1
183
147
Yes

BT-1102
Vip3Ca2
184
105
Yes

BT-1106
Cry42Aa1
185
55
Yes

BT-1110
Cry6Aa3
186
42
No

BT-1113
Cry32Ra1
187
150
Yes

BT-1114
Cry41Aa1
188
96
Yes

BT-1116
Cry15Aa1
189
37
Yes

BT-1120
Cry41Ba2
190
109
Yes

BT-1121
Cry43Ca1
191
137
Yes

BT-1122
Cry64Aa1
192
33
Yes

BT-1124
CryL (BT-0174)
193
119
Yes

BT-1125
Cry73Aa1
194
55
Yes

BT-1126
Cry16Aa1
195
50
Yes

BT-1127
Cry29Aa1
196
61
Yes

BT-1272
Cry66Aa2
197
159
Yes

BT-1274
Vip1Ba1
198
97
Yes

BT-1275
Cry55Aa1
199
44
Yes

BT-1276
Cry55Aa1
200
40
Yes

BT-1277
Cry1Fb4
201
53
Yes

BT-1280
Cry1Ia11
202
83
Yes

BT-1283
Cry12Aa1
203
96
Yes

BT-1284
Cry15Aa1
204
40
Yes

BT-1285
Cry60Ba3
205
35
No

BT-1287
Cry60Aa1
206
35
No

BT-1292
Cry71Aa1
207
71
Yes

BT-1296
Cry22Ab2
208
93
Yes

BT-1297
Cry22Ba1
209
99
No

BT-1300
Cry32Ra1
210
149
No

BT-1305
Cry37Aa1
211
16
Yes

BT-1306
Cry4Ca1
212
60
Yes

BT-1308
Cry59Aa1
213
75
No

BT-1312
Cry45Aa
214
31
Yes

BT-1314
Cry36Aa1
215
57
Yes

BT-1316
Cry49Ab1
216
43
No

BT-1317
Cry36Aa1
217
55
Yes

BT-1318
Cry49Ab1
218
44
No

BT-1319
Cry49Ab1
219
43
Yes

BT-1322
Cry49Aa4
220
29
Yes

BT-1323
Cry4Aa1
221
80
No

BT-1324
Cry4Ba5
222
79
Yes

BT-1331
Cyt2Aa3
223
27
Yes

BT-1333
Cry73Aa1
224
57
Yes

BT-1334
Cry73Aa1
225
57
Yes

BT-1336
Cry64Aa1
226
40
Yes

BT-1337
Cry45Aa
227
41
Yes

BT-1338
Cry45Aa
228
48
Yes

BT-1339
Cry60Aa1
229
38
Yes

BT-1340
Vip1Bb1
230
108
Yes

BT-1342
Vip2Ba1
231
29
No

BT-1343
Vip2Ba2
232
29
No

BT-1344
Vip2Ac1
233
28
No

BT-1345
Vip2Ba2
234
30
Yes

BT-1346
Vip2Aa1
235
33
No

BT-1347
Vip2Ba1
236
30
Yes

BT-1353
Vip2Ba2
237
29
No

BT-1354
Vip2Ac1
238
26
No

BT-1356
Vip2Aa2
239
25
No

BT-1357
Vip2Ba2
240
29
No

BT-1358
Vip2Aa2
241
26
No

BT-1359
Vip2Aa1
242
28
Yes

BT-1360
Vip2Ad1
243
57
No

BT-1362
Vip1Bb1
244
118
Yes

BT-1366
Cry22Aa3
245
75
Yes

BT-1369
Cry3Ca1
246
67
Yes

BT-1371
Cry22Aa3
247
122
Yes

REFERENCES

1. Tokunaga Hi, et al., (2010) Appl Microbiol Biotechnol. 88, 1223

2. Bruce E. Tabashnik, et al., Sci Rep. 2015; 5: 15107

Efficient Protein Expression System for Expressing Bt Insecticidal Protein

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