全 文 ：Received 30 May 2003 Accepted 5 Aug. 2003
Supported by the Knowledge Innovation Program of The Chinese Academy of Sciences (KSCX2-1-01, KSCX2-SW-304), the Hi-Tech
Research and Development (863) Program of China (2002AA211061, 2002AA207003) and National Key Technologies R&D Program in
the 10th Five-Year Plan (2001BA511B03).
* Author for correspondence. E-mail: .
http://www.chineseplantscience.com
Identification and Molecular Mapping of a Stripe Rust Resistance
Gene from a Common Wheat Line Qz180
DENG Zhi-Yong, ZHANG Xiang-Qi*, WANG Xian-Ping, JING Jian-Kang, WANG Dao-Wen
(State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology,
The Chinese Academy of Sciences, Beijing 100101, China)
Abstract: YrQz, a stripe rust disease resistance gene was identified in a common wheat (Triticum
aestivum L.) line Qz180. This resistance was controlled by a single dominant gene, which was confirmed by
genetic analysis of two F2 populations derived from the crosses using Qz180 and two susceptible parents
(Mingxian 169 and WL1). Bulked segregant analysis using simple sequence repeat (SSR) and amplified
fragment length polymorphism (AFLP) markers was conducted in order to map the chromosomal location
of YrQz. The results indicated that YrQz was located on the long arm of wheat chromosome 2B and resided
in a region flanked by two SSR loci Xgwm388 and Xgwm526. Two AFLP markers P35M48(452) and P36M61
(163) were closely linked to YrQz with the genetic distance of 3.4 cM and 4.1 cM, respectively. To our
knowledge, this is the first molecularly mapped stripe rust resistance gene on wheat chromosome 2B.
Key words: wheat; stripe rust disease; disease resistance gene; simple sequence repeat (SSR);
amplified fragment length polymorphism (AFLP)
Stripe rust or yellow rust caused by Puccinia striiformis
f. sp. tritici is one of the most important diseases of wheat
(Triticum aestivum L.) throughout the world. In major wheat-
growing regions in China, stripe rust epidemics often lead
to severe wheat yield losses. The application of fungicides
has shown to be effective in controlling stripe rust to some
extent. However, in the long run, the most economic and
environmentally compatible methods of combating stripe
rust disease are to utilize resistant cultivars in epidemic
regions. Therefore, identification and utilization of stripe
rust-resistant resources are important for improving the
resistance of wheat cultivars. However, because of frequent
emergences of novel stripe rust races, wheat cultivars hav-
ing excellent stripe rust resistance often become suscep-
tible after being grown for some periods of time (Wellings
et al., 1990; Chen et al., 2002). Consequently, the search for
new stripe rust-resistance gene and the breeding of new
resistant wheat varieties should be carried out on a contin-
ued basis. In recent years, marker-assisted selection (MAS)
has been found to be a valuable tool for pyramiding resis-
tance genes, which is generally considered as an effective
way for providing durable protection against plant diseases
(Kumar, 1999). However, the most important requirement
for MAS application is the availability of novel genes with
mapped chromosome positions.
PCR-based markers, such as simple sequence repeat,
also called microsatellite (SSR) and amplified fragment
length polymorphism (AFLP) markers, are currently being
used extensively in genetic mapping and tagging of genes
controlling important agronomic traits in cereal crops (Blair
and McCouch, 1997; Li et al., 2000). In hexaploid wheat,
SSR markers are chromosome-specific, highly polymorphic,
and evenly distributively cover the genome (Roder et al.,
1998; Peng et al., 1999). As a robust technique for DNA
fingerprinting, AFLP is highly informative and well-suited
for mapping of genes with unknown chromosome positions,
particularly in species with low DNA polymorphism (such
as common wheat) (Wang et al., 1997).
In an attempt to find promising resistance source, we
screened a large number of common wheat germplasms with
Puccinia striiformis isolates CY30 and CY31, which are
virulent to majority of Chinese common wheat cultivars. In
this screening, Qz180, which was a land line collected from
Qinghai province, China，was identified as showing ex-
cellent resistance to both of the two isolates, moreover, to
a number of other races including the newest isolate CY32
by the following test (unpublished data). Based on the re-
sistance spectrum, Qz180 was deduced to contain two
known yellow rust (Yr) genes (Yr2, Yr9) and one poten-
tially new Yr gene (tentatively designed as YrQz) with un-
known chromosome position (unpublished data). CY30 or
CY31 was virulent on wheat lines carrying Yr2, Yr9 or both
Acta Botanica Sinica
植 物 学 报 2004, 46 (2): 236-241
DENG Zhi-Yong et al.: Identification and Molecular Mapping of a Stripe Rust Resistance Gene from a Common Wheat Line Qz180 237
as tested (unpublished data). Consequently, YrQz contrib-
uted to Qz180’s excellent resistance. In the present report,
we report the inheritance of YrQz and its chromosome loca-
tion as mapped using SSR and AFLP markers.
1 Materials and Methods
1.1 Plant materials
Common wheat (Triticum aestivum L.) lines, Qz180 and
two susceptible lines Mingxian 169 and WL1, were wheat
germplasm collections in State Key Laboratory of Plant Cell
and Chromosome Engineering, Institute of Genetics and
Developmental Biology, The Chinese Academy of Sciences.
Two F2 populations were constructed by crossing Mingxian
169 and WL1, respectively, with Qz180 as the female parent.
1.2 Resistance tests
Resistance tests were performed on individual F2 prog-
enies and their parents using the pathogenic race CY30.
The primary leaves of F2 seedlings were inoculated with
urediospores. Inoculated plants were grown in the green-
house at 15 ℃ and with 80% humidity. The resistant or
susceptible phenotypes of F2 seedlings were recorded by
comparing to those of their parents. Chi-square tests were
employed to analyze the inheritance of YrQz.
1.3 DNA manipulations
Genomic DNA was extracted from leaf tissues as de-
scribed by Saghai-Maroof et al. (1984). Bulked segregant
analysis (BSA) (Michelmore et al., 1991) was performed
with the bulks consisting of equal amounts of genomic
DNAs from six resistant or six susceptible F2 individuals,
respectively.
1.4 SSR analysis
Approximately 40 ng of genomic DNA was used as the
template for each PCR amplification according to the pa-
rameters established by Roder et al. (1995). After
amplification, 5 mL product was taken from each reaction
and mixed with equal volume of the formamide loading buffer
(98% formamide, 10 mol/L EDTA, 0.1% bromophenol blue,
and 0.1% xylene cyanol). The mixture was electrophoresed
in 6% denaturing polyacrylamide gels containing 6 mol/L
urea. SSR bands were revealed by silver staining as de-
scribed by Bassam et al. (1991).
1.5 AFLP analysis
AFLP experiments were performed followed the proto-
col described by Vos et al. (1995). Genomic DNA was
double-digested with the MseⅠ and PstⅠ at recommended
temperatures. Selective amplification was carried out with
primers carrying three 3-selective nucleotides. The elec-
trophoresis and staining method were the same as those
described in SSR analysis. The denomination for primer
combinations was based on the standard list for AFLP
primer nomenclature①.
1.6 Linkage analysis
The data from SSR and AFLP experiments on individu-
als of F2 populations was analyzed using the Map Man-
ager QTX, a molecular genetic mapping software. The or-
der of the multiple markers was estimated by its “Ripple
function”, and genetic distances were calculated by the
“Kosambi mapping function” with an LOD value > 3.
2 Results
2.1 Inheritance analysis for YrQz
In order to study the inheritance of the new stripe rust
disease resistance gene YrQz, two F2 populations were in-
oculated with CY30 (Table 1). Owing to the apparent
symptom, the resistance or susceptible responses of F2
individuals could be clearly distinguished. For example, in
the population of QZ180× Mingxian 169, the total F2 seed-
lings were divided into 129 resistant and 32 susceptible
individuals according to the infection type 0-0; and 3-4,
respectively, whereas, no seedling with 1-2 scale type was
observed in our test. Chi-square tests showed that the seg-
regation of resistant and susceptible plants in both popu-
lations was consistent with a 3:1 ratio, indicating that YrQz
was inherited as a single dominant gene. The F2 population
with 161 F2 seedlings from the cross Qz180 ×Mingxian
169 was employed in the subsequent SSR and AFLP map-
ping experiments.
2.2 Mapping YrQz with SSR and AFLP markers
In total, 166 wheat SSR primer pairs were chosen for the
mapping experiments. DNA samples from resistance parent
Qz180 and the susceptible line Mingxian 169 were firstly
used as templates for SSR amplification. We found that
Table 1 Segregation of resistant and susceptible progenies in two F2 populations after being inoculated with the wheat stripe rust isolate
CY30
F2 population Plants scored Resistant plants（0-0;） Susceptible plants （3-4） Chi-square tests
Qz180 × Mingxian 169 161 129 32 c23:1 = 1.99Qz180×WL1 89 69 20 c23:1 = 0.180, no visible symptom; 0;, tiny necrotic or chlorotic flecks; 3-4, heavy sporulation.
① Zabeau M, Vos P. 1993. Selective restriction fragment amplification, a general method for DNA fingerprinting. European Patent
Application No. 0534858 A1.
Acta Botanica Sinica 植物学报 Vol.46 No.2 2004238
55% of the primer pairs were polymorphic between the two
parents. Xgwm388 and Xgwm526, located on the long arm
of wheat chromosome 2B (Roder et al., 1998), could reveal
the polymorphisms between parental lines as well as bulks.
A typical amplification pattern generated by Xgwm388 was
shown in Fig.1a. The Xgwm388 allele from the resistant
parent was larger than that from the susceptible parent.
This locus was inherited in a Mendelian co-dominant
manner. There were clear co-segregations between the am-
plification of the larger Xgwm388 allele and the F2 seed-
lings showing the resistant phenotypes (Fig.1a). In the ho-
mozygous susceptible F2 seedlings, only the smaller
Xgwm388 allele was amplified (Fig.1a). In a proportion of
resistant F2 seedlings, both the larger and the smaller alle-
les were amplified, these plants were presumably
heterozygous. In some F2 seedlings, there were discrepan-
cies between the amplification of the alleles and the pheno-
types (Fig.1a, asterisked), these plants were recombinants.
The banding pattern of Xgwm526 alleles was different from
that of Xgwm388 alleles (Fig.1b). Using the primer pair for
Xgwm526, two broad bands (A and B) could be amplified
from the Qz180, whereas, the broad band B and band C
were produced in susceptible parent line Mingxian 169.
Band B did not co-segregate with F2 individuals showing
either resistant or susceptible phenotypes (Fig.1b), so poly-
morphic bands A and C were used to evaluate the linkage
of Xgwm526 with YrQz.
In addition to using known SSR markers, the AFLP/BSA
strategy was also employed to find alternative markers that
may potentially link to YrQz more tightly. In our AFLP
analysis, the enzyme combination of MseⅠ and PstⅠ was
used, and amplification was performed with three selective
nucleotides. Nearly all of the primer pairs could generate
gel patterns with rich and scorable bands and most of them
could display DNA polymorphisms between the two
parents. Approximately 4 000 selectively amplified DNA frag-
ments were generated using 81 primer combinations with
an average of 50 bands amplified for each primer pair. Two
AFLP markers P35M48(452) and P36M61(163) potentially
linked to YrQz were identified. The size of the DNA frag-
ments representing the two markers was 452 and 163 bp,
respectively. As an example, the AFLP pattern of P35M48
(452) in F2 individuals was shown in Fig.2.
2.3 Linkage map construction
According to the preliminary investigation described
above, two SSR loci and two AFLP markers were identified
Fig.1. Amplification of Xgwm388 (a) and Xgwm526 (b) alleles in F2 individuals of Qz180×Mingxian 169. Xgwm526 alleles formed
two band forms, bands A and B in P1, bands B and C in P2. Bands A and C but not B are informative. P1, Qz180; P2, Mingxian 169; R,
resistant progenies; S, susceptible progenies; asterisk, recombinant.
DENG Zhi-Yong et al.: Identification and Molecular Mapping of a Stripe Rust Resistance Gene from a Common Wheat Line Qz180 239
as having a potential linkage relationship with the inter-
ested gene. Then, all of the F2 individuals were analyzed by
these primers to obtain total genotype data, which were
used in our molecular map construction. In order to deter-
mine the linkage pattern among Xgwm388, Xgwm526,
P35M48(452), P36M61(163) and YrQz, the software Map
Manager QTX was applied to establish a multiple point
order of the five chromosomal positions (Fig.3). The result-
ant linkage map showed that YrQz was located in a chromo-
somal region bordered by the two SSR loci Xgwm388 and
Xgwm526 on the long arm of chromosome 2B (Fig.3). The
two AFLP markers P35M48(452) and P36M61(163) flanked
YrQz with genetic distances of 3.4 and 4.1 cM, respectively
(Fig.3) .
3 Discussion
Wheat stripe rust disease is a severe problem for Chi-
nese wheat production all along and has burst on a large
scale more than 10 times in China since the 1950s (Wu and
Niu, 2000). The epidemic was resulted from the breakdown
of the resistance of backbone source due to the occurrence
of new virulent races. Consequently, numbers of cultivars,
which were released or pre-released, would be rendered
susceptible and new virulent races would become the pre-
dominant isolates. In 2002, the latest large epidemic oc-
curred as a consequence of the resistance breakdown of
nearly all Chinese wheat cultivars caused by a new Chi-
nese isolate CY32. At present, the resistant wheat stock for
CY32 is very deficient, so it is a urgent task for wheat ge-
neticist and pathologist to identify, collect and use new
excellent resistance source coming from both common wheat
and wild germplasms. Fortunately, YrQz was identified as
an excellent resistance gene that confers the resistance to a
broad range of isolates including CY32 in our test;
Fig.2. AFLP profile of P35M48(452) in F2 individuals of Qz180×Mingxian 169. Arrow indicates the fragment representing P35M48
(452). P1, Qz180; P2, Mingxian 169; R, resistant progenies; S, susceptible progenies.
Fig.3. Linkage map of YrQz. YrQz was located in a region of
wheat chromosome 2B defined by the two SSR loci Xgwm388
and Xgwm526. The two AFLP markers were more closely linked
to YrQz. Genetic distances in centiMorgans (cM) were calculated
using the Kosambi function.
Acta Botanica Sinica 植物学报 Vol.46 No.2 2004240
furthermore, this promising resistance gene can be easily
incorporated into the elite wheat varieties to enhance their
resistances, since it is contained in common wheat line.
In previous studies on yellow rust disease, a series of
resistance genes, Yr1 to Yr28 as well as many other tempo-
rarily named genes have been identified (Chen et al., 1998).
By using cytogenetic methods, these Yr genes were de-
duced to locate on all wheat chromosomes except 3D and
7A (Chen et al., 1998; McIntosh et al., 1998). Recently,
molecular mapping works for Yr genes have been reported:
using NILs and bulked segregant analysis, Chague et al.
(1999) developed microsatellite and RAPD markers for Yr15
gene of wheat; YrH52, a stripe-rust resistance gene de-
rived from wild emmer wheat, was mapped to chromosome
1B via microsatellite tagging method (Peng et al., 1999); Shi
et al. (2001) identified resistance gene analog polymorphism
(RGA) molecular markers for Yr9; Smith et al. (2002) devel-
oped a sequence-tagged-site (STS) marker for a provision-
ally designated gene YrMoro. Using SSR method, the stripe-
rust gene YrQz from the common wheat line Qz180 was
studied and mapped in a region on the long arm of chromo-
some 2B. This is the first report on molecular mapping of a
Yr gene on this chromosome using SSR markers. However,
owing to the lack of sufficient SSR markers in the region
delimited by Xgwm388 and Xgwm526, we employed AFLP
analysis to find more markers that may potentially more
close linked to YrQz. Since common wheat has a relatively
short evolutional history (Feldman et al., 1995), the genetic
relationship among common wheat varieties is usually very
close. AFLP is a powerful molecular mapping tool and ca-
pable of generating band-rich patterns in between closely
related genotypes. Using AFLP, we indeed obtained two
markers P35M48(452) and P36M61(163) that were consid-
erably closer to YrQz compared to Xgwm388 and Xgwm526.
It has been proposed that, in common wheat, 1 cM genetic
distance is approximately equivalent to 1-3 Mb physical
distances (Delaney et al., 1995). Thus, the deduced physi-
cal distance between the two AFLP markers is at least 7.5
Mb. Because we had not found additional AFLP markers
that are more closely linked to YrQz than P35M48(452) and
P36M61(163), new molecular marker system (such as SNP
markers) will be adopted in the future in order to construct
a higher resolution map of the chromosome region in which
YrQz is resided.
In addition to YrQz, another two Yr genes (Yr5 and Yr7)
have formerly been assigned on the long arm of chromo-
some 2B (McIntosh et al., 1998). At present, we can give
only limited information for the relationships of these genes:
YrQz has a divergent origin from Yr5 that was derived from
Triticum spelta album; Yr7 was originally recorded in the
common wheat cultivar Lee (McIntosh et al., 1998),
however, Yr7 has a different resistant spectrum from YrQz
because it was susceptible to CY30 and CY31 in our test
(unpublished data). To date, there are no molecular map-
ping data for Yr5 and Yr7. Consequently, it is currently
unknown if Yr5 and Yr7 are alleles of YrQz. Therefore, fur-
ther experiments will be carried out to investigate the allelic
relationships among Yr5, Yr7 and YrQz.
In the literatures, there are several successful examples
of converting AFLP markers into sequence characterized
amplified region (SCAR) markers (Shan et al., 1999; Xu et
al., 2001). To utilize YrQz in molecular breeding of resis-
tance to stripe rust, it would be desirable to convert the two
AFLP markers that we identified in this work into SCAR
markers. So experiments are underway to convert P35M48
(452) and P36M61(163) into SCAR markers and to intro-
duce YrQz into elite wheat varieties.
Acknowledgements: The authors are grateful to Profes-
sor WU Li-Ren, Professor WAN An-Min and YANG Dong
(Institute of Plant Protection, The Chinese Academy of
Agricultural Sciences, Beijing, China), and Dr. CAO Zhang-
Jun (College of Agriculture, Northwest Sci-Tech Univer-
sity of Agriculture and Forestry, Yangling, China), for their
excellent assistance in stripe rust disease test.
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