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Weed Science 2007 55:95–101
Genetic Diversity of Wild Oat (Avena fatua) Populations from China and the
United States
Runzhi Li, Shiwen Wang, Liusheng Duan, Zhaohu Li, Michael J. Christoffers, and Lemma W. Mengistu*
Weed genetic diversity is important for understanding the ability of weeds to adapt to different environments and the
impact of herbicide selection on weed populations. Genetic diversity within and among six wild oat populations in China
varying in herbicide selection pressure and one population in North Dakota were surveyed using 64 polymorphic alleles
resulting from 25 microsatellite loci. Mean Nei’s gene diversity (h) for six wild oat populations from China was between
0.17 and 0.21, and total diversity (HT) was 0.23. A greater proportion of this diversity, however, was within (Hs 5 0.19)
rather than among (Gst 5 0.15) populations. For the wild oat population from the United States, h 5 0.24 and HT 5
0.24 were comparable to the values for the six populations from China. Cluster analysis divided the seven populations into
two groups, where one group was the United States population and the other group included the six Chinese populations.
The genetic relationships among six populations from China were weakly correlated with their geographic distribution (r
5 0.22) using the Mantel test. Minimal difference in gene diversity and small genetic distance (Nei’s distance 0.07 or less)
among six populations from China are consistent with wide dispersal of wild oat in the 1980s. Our results indicate that the
wild oat populations in China are genetically diverse at a level similar to North America, and the genetic diversity of wild
oat in the broad spatial scale is not substantially changed by environment, agronomic practices, or herbicide usage.
Nomenclature: Wild oat, Avena fatua L. AVEFA.
Key words: Genetic diversity, gene flow, herbicide selection, simple sequence repeat analysis.
Wild oat is one of the worst annual weeds of the temperate the 2000 collection compared with populations samples in
cereal growing regions of the world. It also is one of the most 1964. For example, resistance frequency increased for
important agronomic weeds in China, particularly as the imazamethabenz from 38% in the 1964 collection to 65%
principal weed in regions growing wheat (Triticum aestivum in the 2000 populations, and for difenzoquat, from 27 to
L.). It was reported that wild oat infested about 5 million ha 58%. Herbicide usage has been affected because of the wild
of cultivated land in China, and wheat yield losses due to wild oat resistance in North America (Mengistu et al. 2003).
oat interference have been conservatively estimated at 1,750 There are no reports in China of herbicide-resistant
million kg of grain annually (Wang et al. 2003). The success biotypes of wild oat so far. However, given the increasingly
of wild oat as a weed depends on long-term seed viability, wide and extensive use of herbicides, e.g., diclofop,
sporadic emergence, production of large numbers of seeds, difenzoquat, fenoxaprop-P, and triallate, especially where
and high genetic variability within populations (Naylor 1983; some of these herbicides have been used for three decades
Naylor and Jana 1976). (Zhang 1995), herbicide-resistance in wild oat may become
Selective herbicides that controlled wild oat, such as one of the most significant weed-management problems
barban, diallate, triallate, and difenzoquat, were introduced facing Chinese growers and researchers in the future.
for wild oat control from the early 1960s and developed into Genetic diversity is the heritable genetic variation within
a large group with different modes of action as reviewed by and among populations of a species. Descriptive studies of
Mengistu et al. (2003). These selective herbicides have been genetic diversity in weed populations can be extremely
used predominantly for wild oat control in North America important because they provide essential background for
and Europe (Cavan et al. 1998; Mengistu et al. 2005). In further focused research, such as understanding the ability of
China, wild oat control practices vary among different regions weeds to adapt to different environments and the impact of
from minimal to high-intensity use of herbicides because of herbicide selection on weed populations. It has been reported
different rural economic circumstances (Wang et al. 2003). that the genetic diversity of wild oat is very high in North
After several decades of frequent and widespread use, wild America and Europe (Cavan et al. 1998; Mengistu et al.
oat has evolved resistance in many countries to several 2005). Furthermore, Mengistu et al. (2005) indicated that
herbicide mechanisms of action (Heap 2006). Wild oat has herbicide-resistant wild oat populations are genetically diverse
developed cross- and multiple-resistance to several herbicide and similar to the herbicide-susceptible populations in North
mechanism-of-action groups, and some populations of wild Dakota. These results suggest that the overall adaptive
oat that had not been exposed to herbicides have even diversity for resistance to new herbicides and other changes
exhibited substantial levels of resistance (Bourgeois et al. in the environment and agronomic practices may not have
1997; Friesen et al. 2000; Rashid et al. 1998; Seefeldt et al. been substantially reduced by herbicide use over the past
1994). Mengistu et al. (2003) studied the herbicide-resistance several decades (Mengistu et al. 2005).
trends among wild oat sampled 36 yr apart. They found that A range of molecular marker systems are used to explore the
the frequency of wild oat resistance was significantly greater in genetic diversity of weeds (Ash et al. 2003; Mengistu et al.
2004, 2005; Mengistu and Messersmith 2002; Moodie et al.
DOI: 10.1614/WS-06-108.1 1997). Among the molecular marker options, microsatellites
* First through fourth authors: Department of Agronomy, China Agricultural are desirable for surveying weed populations because of high
University, Beijing, 100094, China; fifth and sixth authors: Department of Plant
Sciences, North Dakota State University, Fargo, ND 58105. Current address of
levels of polymorphism and information content, selective
sixth author: Minnesota Department of Agricultures, 625 Robert St. N., St. Paul, neutrality, high reproducibility, and wide dispersion in diverse
MN 55155. Corresponding author’s E-mail: lizhaohu@cau.edu.cn genomes (Amsellem et al. 2001; Danquah et al. 2002; Green
et al. 2001; Mörchen et al. 1996; Senda et al. 2004). Although repeat (ISSR) and random amplified polymorphic DNA
there are a few reports on the development of microsatellite (RAPD) markers (Cavan et al. 1998; Mengistu et al. 2005).
markers in oat (Avena sativa L.) (Li et al. 2000; Narinder et al. This information can improve our understanding of the
2002), this technique has not been used for analysis of genetic genetic diversity of wild oat from a broad spatial scale within
structure and population differentiation in wild oat. Further- China and between two continents and can assist in
more, there are similarities in genome organization and maintaining an effective management strategy for wild oat
nucleotide sequences of genes among grass species (Devos and control with increasing herbicide usage in China.
Gale 2000), and information gained during analysis of rice
(Oryza sativa L.), wheat, and barley (Hordeum vulgare L.)
should apply to the study and manipulation of genomes of Materials and Methods
relatively less-well-studied species like wild oat. Material Collection and DNA Extraction. A total of 1,000
We assume the genetic diversity of wild oat in China is high individual wild oat plants, representing six populations, were
and comparable to the genetic diversity of wild oat in North collected in 2004 from six provinces in China, where wild oat
America and Europe, as reported by Mengistu et al. (2005) is one of the worst weeds of different farming systems from
and Cavan et al. (1998). However, the genetic variation of winter wheat to spring wheat and where herbicide-selection
wild oat in China has never been investigated on a large pressure differs from minimal to heavy use because of regional
geographic scale, and detailed information on the genetic economic status (Figure 1, Table 1). The sampled popula-
structure of wild oat populations has remained unavailable. tions inhabited different areas within a wide geographic range
Here, we report a study using microsatellite markers to detect (100 to 119uE, 32 to 41uN), and each population was
genetic diversity within and among six populations of wild oat separated by at least 180 km.
sampled across a large spatial range in China, where farming Wild oat samples were collected as described by Mengistu
systems are substantially different from winter wheat to spring et al. (2003). Each population was created by collecting three
wheat production areas, and herbicide-selection intensity is to five samples per site, moving 1.5 to 2 km between
significantly different (from minimal to heavy use). One sampling sites. Seeds for each sample were from a single
population from North Dakota, where wild oat control is mature parent plant, and the location of each plant was
predominantly by herbicides (Mengistu et al. 2003), is documented using a hand-held Global Positioning System
included for comparison. (GPS). If wild oat was not present at the target sampling site,
Our objectives were (1) to evaluate the usability of oat the next occurrence of wild oat was sampled. One plant of the
microsatellites and validate whether barley and wheat three to five samples per site was randomly selected for further
microsatellites could be used to study the genetic diversity study. Thus, of the 1,000 plants sampled, the analysis used
of wild oat, (2) to determine the genetic diversity and 347 samples representing six populations (provinces), with
differentiation of seven wild oat populations, and (3) to a sample size of 41 to 83 plants per population. In addition,
compare the levels of genetic diversity detected with this study included 68 individual plants of one wild oat
microsatellites to those found with intersimple sequence population from North Dakota, where wild oat control is
predominantly by herbicides, as described by Mengistu et al. PCR programs used for different microsatellite primers
(2003). were followed: from Li et al. (2000), for PCR program 1, 2,
Seeds from each sample were sown in the greenhouse and and 3; from Narinder et al. (2002), for program 4; and from
grown for 3 wk. The newly expanded leaf from each plant was Guyomarc’h et al. (2002), for program 5 (Table 2). PCR
frozen in liquid nitrogen and ground in a microcentrifuge products were separated on a sequencing gel containing 6%
tube. DNA was extracted following a rapid one-tube polyacrylamide, 7 M urea, and 13 TBE at 85 W constant
extraction (ROSE) method (Steiner et al. 1995). power for 45 min (BioRad sequencing system).2 The gel was
fixed, stained, and dried using a DNA silver-staining
Microsatellite Amplification. Optimal microsatellite primers technique (Bassam et al. 1991).
were selected by screening 45 oat primers (Li et al. 2000;
Narinder et al. 2002), 38 barley primers (Liu et al. 1996), and Statistical Analysis. Microsatellites are codominant genetic
294 wheat primers (Guyomarc’h et al. 2002). Among 377 markers, which can distinguish heterozygotes from homo-
microsatellite primers, seven oat primers (nine loci), seven zygotes. In this study, because wild oat is predominantly self-
barley primers (nine loci), and six wheat primers (seven loci) pollinated, with few heterozygous plants, and null alleles (i.e.,
produced clear and simple reproducible fragments and were alleles that do not give a PCR product) were observed using
used to amplify 415 wild oat individuals (Table 2). barley and wheat microsatellites in our study, we viewed these
The reaction proceeded on a GeneAmp PCR system 9700 markers as dominant markers in our data analysis. Only the
(PE Biosystems).1 Microsatellite amplifications were per- number of alleles per locus was calculated. Genetic diversity
formed in a 20-ml volume, containing 40-ng template DNA, parameters were calculated using 64 alleles from 25 micro-
13 polymerase chain reaction (PCR) buffer (10 mM Tris– satellite loci.
HCl, pH 8.5, 1.5 mM MgCl2, 50 mM KCl, 0.001% Nei’s (1973) gene diversity (h), Shannon’s information
gelatin), 200 mM of each deoxynucleoside-59-detriphosphate index (I ) (Lewontin 1972), total diversity (HT), diversity
(dNTP), 0.2 mM of each forward and reverse primer, and 1 U within (HS) and among populations (GST), gene flow among
Taq DNA polymerase. populations (Nm), and Nei’s (1978) unbiased genetic identity
Table 2. Microsatellite primer sequences, their PCR programs, and number of alleles detected.a
Primer name Primer sequence (59–39) PCR Alleles Primer name Primer sequence (59–39) PCR Alleles
Oat SSRs Barley SSRs
AM6 GGATCCTCCACGCTGTTGA 2 2 HVM4 AGAGCAACTACCAGTCCAATGGCA 1 2
CTCATCCGTATGGGCTTTA GTCGAAGGAGAAGCGGCCCTGGTA
AM11 TCGTGGCAGAGAATCAAAGACAC 1 2 HVM13 AGTAGCTATGTGTTTGGATCGC 3 2
TGGGTGGAGGCAAAAACAAAAC CATCAAGGGCATCCTCATG
AM14 GTGGTGGGCACGGTATCA 2 3/4b HVM30 AGTGGGGAATGAGAGAATGG 3 2/2
TGGGTGGCGAAGCGAATC TGCTTGTGGGTCATCACAC
AM31 GCAAAGGCCATATGGTGAGAA 2 6 HVM51 TCTAAATTACCTTCCCAGCCA 3 2
CATAGGTTTGCCATTCGTGGT AAGCAGACATGTAGGAGGTCA
AM42 GCTTCCCGCAAATCATCAT 2 3 HVM64 GATGTGAAGGCTGCCTG 3 1
GAGTAAGCAAAGGCCAAAAAGT ACACGCCCTATTACCCAGTG
AM102 TGGTCAGCAAGCATCACAAT 4 3 HVM67 GTCGGGCTCCATTGCTCT 3 2/2
TGTGCATGCATCTGTGCTTA CCGGTACCCAGTGACGAC
AM112 AGCGGTGTAGGGGAAAGAGT 4 12/1 HVM68 AGGACCGGATGTTCATAACG 3 2
TTCTTGGTTTAGATGGGAGGA CAAATCTTCCAGCGAGGCT
Wheat SSRs Wheat SSRs
Ba119 CACCCGATGATGAAAAT 5 1 Ba148 GCGCAACCACAATGTATGCT 5 2
GATGGCACAAGAAATGAT GGGGTGTTTTCCTATTTCTT
Ba120 CCCCCTCTCTTCCTCAT 5 2 CFD-8 ACCACCGTCATGTCACTGAG 5 1
ATATAGCTCCCCCATTTCCT GTGAAGACGACAAGACGCAA
Ba128 GCGGGTAGCATTTATGTTGA 5 1 CFD-13 CCACTAACCAAGCTGCCATT 5 2/2
CAAACCAGGCAAGAGTCTGA TTTTTGGCATTGATCTGCTG
a
Abbreviations: PCR, polymerase chain reaction; SSR, simple sequence repeat.
b
The number of alleles detected from two loci of one primer.
HT 5 0.36 for five wild oat populations and 0.31 for five Nei’s unbiased genetic distance among six wild oat
animated oat (Avena sterilis L.) populations (Cavan et al. populations from China ranged from the least at 0.006
1998) but is comparable with HT values for other self- between the Anhui and Henan populations to the largest at
pollinating weeds, such as 0.21 for blackgrass (Alopecurus 0.07 between the Jiangsu and Inner Mongolia populations
myosuroides Huds.) (Chauvel et al. 1994) and 0.25 for annual (Table 4). Nei’s genetic distance between the U.S. population
bluegrass (Poa annua L.) (Mengistu et al. 2000). The reason and the six Chinese populations ranged from 0.09 to 0.14. Of
for this difference can partly be explained by the limited the six Chinese wild oat populations, the Qinghai population
number of samples from the United States in our study and shared the most genetic identity with the U.S. population,
a limited number of suitable microsatellite markers. with a Nei genetic identity of 91% and a Nei genetic distance
of 0.09.
Genetic Structure and Deviation of Wild Oat Populations. Wild oat in the 1970s was mainly distributed in Qinghai
Most of the total genetic diversity of wild oat in China existed and Inner Mongolia provinces, and it had begun to invade the
within populations (HS 5 0.19), whereas only a small amount Shaanxi, Henan, Anhui, and Jiangsu provinces by the early
of genetic diversity (15%, GST 5 0.15) was present among 1980s and has become one of the worst weeds of these
populations (Table 3). The level and distribution of genetic agricultural fields (Tu 1987). Broad dispersal of wild oat in
the past two decades has contributed to the small genetic
diversity in plant species is affected by the mating system.
distance (0.07 or less) among populations.
Compared with self-pollinated species, population differenti-
The six Chinese wild oat populations formed two groups of
ation in cross-pollinated species is generally less apparent and
clusters in an unweighted pair group method with arithmetic
much of the variation characteristic of the species may be
mean (UPGMA) dendrogram, whereas the United States
found within a population (Hamrick and Godt 1990; population was very different (Figure 2). One main group was
Loveless and Hamrick 1984). Wild oat is predominantly composed of the Anhui, Henan, Jiangsu, and Qinghai
self-pollinated, but the results of the present study indicate populations, and the Shaanxi and Inner Mongolia populations
85% of the total genetic diversity was within populations formed the other group. Within the first group, the Anhui
(Table 3). and Henan populations were one subgroup and were more
One possible explanation for the high within-population similar to each other than within clusters of the other
diversity is that each population covered a relatively large area. populations.
The samples for every population used for this study account A Mantel test to compare matrices of genetic and
for just 30% of original plant samples collected. For example, geographic distances did not show a relationship between
the original materials of the Qinghai population included 230 them (r 5 0.22). For example, the four populations in the
samples, but only 67 were used, and those were distributed first cluster had two populations sampled from sites closest to
between the extreme west (100u579E) to extreme east each other, Anhui and Henan (180 km), and also two
(102u489E) and the extreme north (36u429N) to extreme populations that were farthest from each other, Qinghai and
south (36u199N). Thus, these samples for every population Jiangsu (1,680 km) (Figures 1 and 2). Interestingly, the
represented specific and abundant variation that likely Henan and Anhui populations were comparatively close in
remained within populations. In addition, recent evidence distance to the Shaanxi population (450 km and 620 km,
has revealed high levels of intrapopulation variation in self- respectively) whereas the Inner Mongolia population was very
pollinated plants. For example, Mengistu, et al. (2005) found far from the Shaanxi population (810 km). However, the
that between 9% and 16% of total diversity remained within Shaanxi and Inner Mongolia populations formed one group
wild oat populations. and had 95% microsatellite identity, whereas the Henan and
Based on an indirect estimate of gene flow, where Nm 5 Anhui populations belonged to another group.
[0.25(1 2 GST)/GST] (Slatkin and Barton 1989), gene flow Our study revealed that the genetic diversity of wild oat
among the six Chinese wild oat populations was high (Nm 5 populations in China was high and comparable with the status
1.4) (Table 3). Genetic drift will result in substantial local of the U.S. population. Weed species with high levels of
differentiation if Nm , 1 but not if Nm . 1 (Slatkin 1987). genetic diversity exhibit considerable potential for weed
The Henan, Anhui, and Jiangsu provinces are the main wheat adaptation and, therefore, the effectiveness of frequently used
production areas of China, and there has been frequent wheat- weed-control techniques may be reduced (Dekker 1997; Holt
seed transport with other provinces, such as Qinghai, Inner and Hochberg 1997). However, there aren’t any reports of
Mongolia, and Shaanxi. Thus, seed migration is one possible herbicide-resistant biotypes of wild oat in China, whereas the
mechanism of high gene flow among populations. U.S. wild oat populations have evolved resistance to a wide
Li et al.: Genetic diversity of wild oat from China and U.S. N 101