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Bioscience Methods
BM 2010, Vol.1, No.1
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Research Article Open Access
Developing Specific Markers and Improving Genetic Mapping for a Major
Locus
Tyr1
of Citrus Nematode Resistance
Xiang Xu
1
, Deng Zhan'ao
2
, Zheng Qifa
3
, Chen Cunxian
3
, Frederick G. Gmitter Jr.
3
1 Institute of Fruit Tree Research, GDAAS, Guangzhou, 510640; 2 University of Florida, GCREC, Bradenton, FL34203, USA; 3 University of Florida,
CREC, Lake Alfred, FL33850, USA
Corresponding author email: fgg@lal.ufl.edu;
Authors
Bioscience Methods 2010, Vol.1 No.1 DOI:10.5376/bm.2010.01.0001
Received: 2 Apr., 2009
Accepted: 29 Apr., 2009
Published: 28 May, 2009
This article was first published in the Molecular Plant Breeding (Regular Print Version), and here was authorized to redistribute 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 as:
Xu et al, 2009, Developing Specific Markers and Improving Genetic Mapping for a Major Locus
Tyr1
of Citrus Nematode Resistance, Molecular Plant
Breeding, 7(3): 497-504
Abstract
The NBS-LRR class resistance-gene candidate sequences (Pt8a and Pt9a) were used to develop new specific markers
to the citrus nematode resistance gene locus
Tyr1
. By high-density colony screening, over 200 positive clones were pulled out from
the BAC library. A few of the clones were found to be closely linked with the
Tyr1
region, because the primers from these clones
insert sequence produced polymorphism which matched up with the phenotype after bulked segregant analysis. By primer walking
approach, three integrate NBS-LRR class resistance-gene sequences were tagged and identified separately in three clones (7A4,
4L17 and 29F20). More specific markers were developed from these tagged sequences and relatively high-density genetic maps
were constructed by incorporating the newly developed markers and previously developed markers in the ‘9145 family’. New
markers were applied in ‘9401 family’ trying to estimate roughly the genetic distance between the Ctv and Tyr1 region.
Keywords
Citrus; Nematode; Resistance gene; Molecular marker; Genetic map
Background
Citrus nematode [CN,
Tylenchulus semipenetrans
(Cobb 1914)] has been one of the major parasitic
pests that cause loss in production and a reduction
in quality of fruit throughout the world (Duncan and
Cohn, 1990). Citrus rootstock breeding for
nematode resistance is also one of the objectives of
modern citrus genetic research and molecular
engineering. Although the inheritance of citrus
nematode resistance is not yet clear, the QTLs of
the citrus nematode resistance had been mapped
with RAPD markers indicating a major gene confers
resistance to citrus nematode (Ling et al., 2000).
The construction of high-density genetic map is
very important for tagging the genes or QTLs, as
well as for map-based cloning (MBC) and
marker-assisted selection (MAS). The development
of DNA markers has resulted in the genetic linkage
map construction in most crop species, including
perennial crop of citrus. Several loci of horticultural
significance in citrus had been tagged with RAPDs,
SCARs, CAPS and other markers, such as the citrus
tristeza resistance gene (
Ctv
) and the major gene for
citrus nematode resistance (
Tyr1
) (Gmitter et al.,
1996; Deng et al., 1997; 2000; Fang et al., 1998;
Ling et al., 2000). In recent years, the way of
resistance-gene analogs (RGAs) or resistancegene
candidates (RGCs) analysis by degenerate primers
has been quickly and broadly extended. The
NBS-LRR (nucleotide-binding site, Leucine-rich
repeat) class of resistance genes have been proved
to be most prevalently existed (Shen et al., 1998;
Speulman et al., 1998; Meyers et al., 1999; Deng et
al., 2000; Noir et al., 2001; Madsen et al., 2003; Lee
et al., 2003). These genes share striking structural
similarities, which seems to encode receptors that
detect the presence of specific pathogens, and
suggests that certain signaling reaction are common
to all or most plant defense systems (Lee et al.,
2003). Ten classes of NBS-LRR sequences were
found in citrus with RGC-derived primers (Deng et
al., 2000). Three of the RGC-derived markers