3 Biotech (2018) 8:252
Construction of a synthetic Araneus ventricosus dragline silk gene
multimer and its expression in Escherichia coli
· Anwen Liang
· Ziqiang Liang
· Guanghong Li
· Fanghai Wang
Received: 2 August 2017 / Accepted: 7 May 2018 / Published online: 11 May 2018
© Springer-Verlag GmbH Germany, part of Springer Nature 2018
One of the most representative core gene sequence of Araneus ventricosus dragline silk protein partial cDNA monomer
(JN857964.2) was selected and multimerized using a “head-to-tail” strategy by compatible but nonregenerable sites at both
ends resulting in a concatemer of 16 contiguous monomers. This concatemer was cloned into pET-28a(+) expression vector
and transformed into Escherichia coli. A 52.6 kDa silk protein was successfully expressed and detected by SDS–PAGE and
conﬁrmed by Western blotting. A maximum yield of the silk protein was expressed with 7.06 mM IPTG after 5 h incubation.
This is the ﬁrst report on the construction and overexpression of a A. ventricosus dragline silk multimeric gene construct and
the results from our study will provide a reference point for further exploration and development of large-scale production
of spider silk protein.
Keywords Araneus ventricosus · Spider dragline silk protein · Multimer · Expression
Spider dragline silk is a protein ﬁber, which possess numer-
ous excellent characteristics, such as, extreme lightness and
softness, high strength and extensibility, good toughness
and elasticity, strong shock resistance ability, resistance to
low temperature, biodegradability, etc. Due to these unique
properties, spider dragline silk has numerous applications.
In military, it can be used to make bullet-proof vests, para-
chutes, armors in aircrafts and tanks. In medicine, it is useful
in manufacturing artiﬁcial joints and limbs, as sutures for
stitching wounds, as scaﬀolds for cell culture, and many bio-
logical tissue materials. Industrial applications of dragline
silk include manufacturing of wheels, tyres and ﬁshing nets.
In architecture, it may be used as structural and composites
for buildings and in textile manufacturing it can be used to
make clothes, scarves, hats, etc. (Pan et al 2004; Zhao et al
Spiders spin a very small amount of silk, and the process
of silk extraction from spiders is complex. Additionally,
spiders can not be reared in high-density farming due to their
cannibalistic and aggressive nature and hence is diﬃcult to
obtain suﬃcient quantities of natural spider dragline silk to
satisfy the demands for practical applications (Wang and Hu
2007). Therefore, research has focused on producing spider
silk using alternate bioengineering technology. A spider
dragline silk protein gene from Nephila clavipes, has been
successfully cloned and expressed in Escherichia coli (Fah-
nestock and Irwin 1997; Xia et al. 2010; Qiao et al. 2014)
and Saccharomyces cerevisiae (Fahnestock and Bedzyk
1997). Several other recombinant spider silk proteins have
been produced from many species using genetic engineering
strategies, including Argiope aurantia, Nephilengys cruen-
tata, Parawixia bistriata and Avicularia juruensis (Tokareva
et al. 2013).
We report in this paper for the ﬁrst time cloning and over-
expression of a spider silk protein from Araneus ventricosus.
A. ventricosus is one of the common spiders in China, which
has much larger webs and a better spider silk compared to
other kind of spider silks.
* Fanghai Wang
State Key Laboratory for Biocontrol and Institute
of Entomology, Sun Yat-sen University, Guangzhou 510275,