Molecular Pathogens 2012, Vol.3, No.3, 12
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18
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12
Research Report Open Access
Molecular Identification and Sequence Analysis of the Apple Scar Skin Viroid
(
ASSVd) Isolated from Four Kinds of Fruit Trees in Xinjiang Province, China
Yuting Wang , Ying Zhao , Jinxin Niu
Horticultural Department, Agricultural College of Shihezi University, Shihezi, 832003, P. R., China
Corresponding authors email:
njx105@163.com;
Authors
Molecular Pathogens, 2012, Vol.3, No.2 doi: 10.5376/mp.2012.03.0002
Received: 26 Jul, 2012
Accepted: 20 Aug, 2012
Published: 23 Aug, 2012
This is an open access article published 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:
Wang et al., 2012, Molecular Identification and Sequence Analysis of the Apple Scar Skin Viroid (ASSVd) Isolated from Four Kinds of Fruit Trees in Xinjiang
Province, China, Molecular Pathogens, Vol.3, No.3 12-18 (doi: 10.5376/mp.2012.03.0003)
Abstract
In this paper, low molecular weight RNAs were extracted from tender leaves and shoots of apple, pear, peach and apricot
trees, which were collected from Yanqi, Hejing, Bohu, Heshuo, Xinhe, Korla and Aksu in Xinjiang province, China. Then the
samples were detected by reverse transcription-polymerase chain reaction (RT-PCR) and
in situ
RT-PCR technology. The detection
results demonstrated that four kinds of fruit trees were infected by
Apple scar skin viroid
(
ASSVd) and the detection rates were 2.1%,
2.1%, 2.8%
and 3.3% on apple, pear, peach and apricot trees, respectively.
In situ
RT-PCR result further confirmed that the existence
of ASSVd was mainly distributed in the nucleus of the leaf tissues. And then the RT-PCR products from the samples were cloned and
sequenced, and we obtained 42 ASSVd nucleic acid sequences, which were registered in GenBank and the accession numbers was
from EU031455 to EU031496 by using biological software to analyze and the total detection rate was 3.0%. The homology analysis
results showed that the 42 isolated ASSVd sequences had 85%~100% nucleotide sequence identity with previously published
sequence NC_001340 (Puchta et al.,
1990).
All this results indicated that the variation of nucleic acid sequence of ASSVd isolate in
each host was not obvious and was no significant difference in regions and varieties. In this present study, we established the
optimized detection methods of RT-PCR and
in situ
RT-PCR, which would lay a good foundation for the rapid identification of
ASSVd in these four kinds of fruit trees.
Keywords
Apple tree; Pear tree; Peach tree; Apricot tree;
Apple scar skin viroid
(
ASSVd); RT-PCR;
In situ
RT-PCR; Sequence analysis
Introduction
Apple scar skin viroid
(
ASSVd), belongs to
Apscaviroid
,
usually contains 330 nucleotides with
the central conserved region but no enzyme activity,
forming an asymmetrical rolling loop replication (Li
and Sano, 2000). ASSVd was firstly reported to
infect pome fruit trees (
Malus
,
Pyrus
and
Cydonia
spp.)
(
Hashimoto and Koganezawa, 1982). Subsequently,
Chen et al (1986) also detected the viroid from the
shoots of disease trees, cloned and analyzed its
nucleic acid sequence and demonstrated its
pathogenicity of this viroid, and then people have
confirmed that apple scar skin disease is a disease
cause of viroid (Zhou et al., 2002). Guo et al (2005)
have cloned and analyzed the sequence of ASSVd in
Liaoning province, China.
ASSVd mainly infected apple trees and pear trees
(
Hadidi et al., 1991; Yang et al., 1992; Koganezawa
et al., 2003; Kyriakopoulou et al., 2003). Because the
viroid was replicated and accumulated in nucleus of
it’s hosts, it can be detected in the leaf, stem, epidermis
and rootstock of the hosts disease-susceptibility, as well
as the epidermis, pulp and seed of the fruits (Hadidi et
al., 1991). Consequently, the fruit tree which was
infected with the viroid would carry it all its life, and
it can be spread via grafting and prune tools.
In this study, we detected and amplified 42 ASSVd
isolates, and then the full length sequences of the 42
ASSVd were sequenced from apple, pear, apricot and
peach trees in Xinjiang of China. The distribution of
ASSVd was confirmed by
in situ
RT-PCR method.
Thus, the results of our study also showed that ASSVd
was a pathogen of peach and apricot trees. Meanwhile,
we established the optimized detection methods of
RT-PCR and
in situ
RT-PCR, which would provide a
basis for the rapid identification of ASSVd.
