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Bt Research (Online) 2010, Vol.1 No.3
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Research Article Open Access
Expression and Localization of Cry1Ac22 Crystal Protein from
Bacillus
thuringiensis
W015
-
1 in Yeast (
Saccharomyces cerevisiae
)
Shenkui Liu
1*
, Zhuoming Liu
2,3*
, Youzhi Li
3
, Xuanjun Fang
1,2,3
1. Alkali Soil Natural Environmental Science Center (ASNESC), Northeast Forestry University, Harbin, 150040, P.R. China
2. Haide Institute of Tropical Agricultural Resources (HITAR), Sanya, 572025, P.R. China
3. College of Life and Technology Science, Guangxi University, Nanning, 530004, P.R. China
* These authors contributed equally
Corresponding author, xuanjunfang@hitar.org;
Authors
Bt Research 2010, Vol 1 No 3
DOI: 10.5376/bt.2010.01.0003
Received: 27 Aug., 2010
Accepted: 21 Dec., 2010
Published: 30 Dec., 2010
This is an Open Access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and
reproduction in any medium, provided the original work is properly cited.
Preferred citation for this article:
Liu et al., 2010, Expression and Localization of Cry1Ac22 Crystal Protein from
Bacillus thuringiensis
W015-1 in Yeast (
Saccharomyces cerevisiae
), Bt
Research (online), Vol.1 No.3 (DOI: 10.5376/bt.2010.01.0003)
Abstract
In order to explore the expression and sub-cellular localization of
Bt
Cry1Ac22 insecticidal crystal protein in
eukaryotic organism, we constructed eukaryotic expression vector pYES2
-
Cry1Ac22
-
GFP by fusing Cry1Ac22 and green
fluorescent protein (GFP) based on the initiative pYES2 vector. The construct was transformed into yeast (
Saccharomyces cerevisiae
)
strain INVScl to express the fusion gene promoted by GAL1. With the induction of 20%galactose, the fusion protein of
Cry1Ac22-GFP was expressed as a mass of fluorescence particles in the yeast cell. SDS-PAGE analysis showed that the fusion
proteins were expressed in the yeast crude succus about 130 kD in size. The fusion proteins can generate intense fluorecense light
alone with the yeast cell membrane observed under the fluorescent microscope, which indicated that Cry1Ac22 proteins were
anchored in the cell membrane. This research might provide the insights and approach for studying the function of
Bt
insecticidal
proteins as well as for tracing the trail of transgenic proteins.
Keywords
Bt
(
Bacillus thuringiensis
); Yeast (
Saccharomyces cerevisiae
); Cry1Ac22; Green fluorescent protein (GFP);
Eukaryotic expression; Subcellular localization
Background
Bacillus thuringiensis
(
Bt
) is a ubiquitous gram-
positive bacterium, which produces parasporal
crystal protein during sporulation that has insecticidal
action (δ
-
endotoxin). It attracts increasing attention
due to its insecticidal properties to agricultural
pests(Li et al., 1991; Knowles and Ellar, 1986).
Bt
toxin Cry1Ac22 insecticidal crystal proteins are
isolated from
Bacillus thuringiensis
W015
-
1 in the
intestines of diapausing larvae of silkworm
(
Bombyx mori
). It is known that
Bt
W015
-
1 has
higher insecticidal activity than HD73 on some
lepidopteran insects such as
Clanis bilineata Walker
,
Helicoverpa armigera
,
Spodoptera litura
,
Plutella
xylostella
(Xie et al., 2010).
Cry1Ac22 insecticidal crystal protein, encoded by
cry1Ac22
gene, is constituted by 1 178 amino acids,
It is different from Cry1Ac1 protein (isolated from
HD73 (Adang et al., 1985)) in amio acid sequence
at sites of 233 (T/R), 448 (M/I) and 1158 (K/E) (Xie
et al., 2010).
cry1Ac22
gene can express 133 kD
proteins in
Escherichia coli
, and the purified
proteins show high insecticidal activity to
Plutella
xylostella
(Xie et al., 2010; Liu et al., 2010). Taking
into account that W015
-
1 was isolated from the
intestines of diapausing larvae of silkworm and its
cry
gene shows apparent diversity with HD73, the
strain and its insecticidal proteins could be used as
candidate strain for integrated control and resistance