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Genomics and Applied Biology, 2011, Vol.2 No.1
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Research Article Open Access
Cloning of an Ascorbate Peroxidase Gene from
Puccinellia tenuiflora
and its
Expression Analysis
Qingjie Guan
1,2
Lin Li
1
Takano Tetsuo
2
Shenkui Liu
1
1. Alkali Soil Natural Environmental Science Center (ASNESC), Northeast Forestry University, Harbin, 150040; 2. Asian Natural Environment Science
Center (ANESC), The University of Tokyo, Tokyo, 1880002, Japan
Corresponding author, shenkuiliu@nefu.edu.cn;
Authors
Genomics and Applied Biology 2011, Vol.2 No.1 doi: 10.5376/gab.2011.02.0001
Received: 10 Sep., 2010
Accepted: 11 Feb., 2011
Published: 25 Feb., 2011
This article was first published in Genomics and Applied Biology (Regular Print Version) in Chinese, and here was authorized to redistribute in English 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:
Guan et al., 2009, Cloning of an Ascorbate Peroxidase Gene from Puccinellia tenuiflora and its Expression Analysis, Genomics and Applied Biology, 28(4):
631-639 (doi: 10.3969/gab.028.000639)
Abstract
The full-length gene of an ascorbate peroxidase (PutAPx) was isolated from
Puccinellia tenuiflora
OhwicDNA library.
The gene is 1 125 bp in length and it has an ORF of 876 bp, which encoded a protein of 291 amino-acid with an estimated molecular
weight 32 kD and an isoelectric point of 7.71. Blasting at NCBI, we found that
PutAPx
showed high similarity (89.7%, 94.3%,
94.2%, 50.7% and 79.6%) to 5 different gramineae species (
Oryza sativa
L.,
Hordeum vulgare
L.,
Triticum aestivum
,
Lolium perenne
L., and
Zea mays
). The result of phylogenetic tree showed that PutAPx has the closdest genetic distance with
Hordeum vulgare
L.
and
Triticum aestivum
. Transgenic yeast (InVSC1), expressing
PutAPx
gene, under the inducement with
β
-galactose showed higher
stress resistance in oxidation stress than control. In this study, we successfully cloned an ascorbate peroxidase (PutAPx) and studied
in primary levle, which laid the foundation for the future study in the mechanism of oxidative stress mechanism of the role of the
foundation establish.
Keywords
Puccinellia tenuiflora Chinampoensis
Ohwi; Ascorbate peroxidase; Gene cloning
Background
Puccinellia tenuiflora Chinampoensis Ohwi
which
grows in meadow steppe, saliferous soil of North
China, is a perennial gramineous herbage, with strong
resistance to saline-alkali. Because of long-term
evolution and selection, the young seedlings with only
five leaves can grow well in soil where alkalinity
exceeds pH 10 and the salt content of surface soil
overrides 5% (Li and Yang, 2004). Therefore,
Puccinellia tenuiflora is not only a superior herbage,
but also a precious halobiotic germplasm resource,
saline-alkali-tolerant gene resource.
Nowadays,
isolating saline-alkali-resistant gene from
halophytes
are
the mechanism of plant
resistance to
saline-alkali and the hinge of molecular
selective
breeding
saline-alkali-resistant cultivars.
Puccinellia
tenuiflora
is now paid widely attention by the
researchers who engage in saline-alkali-stress resistance
and its genes also have been cloned and published on
NCBI GenBank database one after another, including
betaine aldehyde dehydrogenase (EF095710), EF095710),
H
+
-ATPase (DQ090006), NADH-glutamine synthetic
protein (DQ093360), heatshock protein (DQ093361),
glutathione transferase (DQ093362), Na
+
/H
+
pump
(EF440291), H
+
pump PutCAX1 (AB472071), ferritin
(DQ090999), PutPMP3
-
1(AB363567), PutPMP3
-
2
(AB363568), Actin (FJ545 641), Put-R40g3 (AB465547),
dehydroascorbate reductase protein (DQ090998),
Put-Cu/Zn-SOD and so on. Under salt stress, cell
could produce active oxygen (ROS) such as
oxyradical (O
2-
), hydrogen peroxide (H
2
O
2
), hydroxyl
radical (OH
-
), which could cause the oxidative stress
(Shan et al., 2006). With high specificity and affinity
to ascorbic acid, APX (ascorbate peroxidase, APX,
EC1.11.1.11), which is the main enzymes for
eliminating H
2
O
2
, catalyzing H
2
O
2
to reduce into H
2
O
by the reductive ascorbic acid substrate, produces
dehydroascorbic acid, and the acid can be reduced to
ascorbic acid through many different pathways
coupling with H
2
O
2
consumption (Asada, 1992;