Multiple allelic forms of acetohydroxyacid synthase are responsible for herbicide resistance in <Emphasis Type="Italic">Setaria viridis</Emphasis>

In weed species, resistance to herbicides inhibiting acetohydroxyacid synthase (AHAS) is often conferred by genetic mutations at one of six codons in the AHAS gene. These mutations provide plants with various levels of resistance to different chemical classes of AHAS inhibitors. Five green foxtail [Setaria viridis (L.) Beauv.] populations were reported in Ontario with potential resistance to the AHAS-inhibiting herbicide imazethapyr. The objectives of this study were to confirm resistance, establish the resistance spectrum for each of the five populations, and determine its genetic basis. Dose response curves were generated for whole plant growth and enzyme activity, and the AHAS gene was sequenced. Resistance was confirmed by determining the resistance factor to imazethapyr in the five resistant green foxtail populations for whole plant dose response experiments (21- to 182-fold) and enzyme assays (15- to 260-fold). All five imazethapyr-resistant populations showed cross-resistance to nicosulfuron and flucarbazone while only three populations had cross-resistance to pyrithiobac. Sequence analyses revealed single base-pair mutations in the resistant populations of green foxtail. These mutations were coded for Thr, Asn, or Ile substitution at Ser₆₅₃. In addition, a new mutation was found in one population that coded for an Asp substitution at Gly₆₅₄. There is an agreement between the spectra of resistance observed and the type of resistance known to be conferred by these substitutions. Moreover, it indicates that, under similar selection pressure (imazethapyr), a variety of mutations can be selected for different populations, making the resistance pattern difficult to predict from herbicide exposure history.     

An isoleucine-leucine substitution in chloroplastic acetyl-CoA carboxylase from green foxtail (<Emphasis Type="Italic">Setaria viridis</Emphasis> L. Beauv.) is responsible for resistance to the cyclohexanedione herbicide sethoxydim

The cDNAs encoding chloroplastic acetyl-CoA carboxylase (ACCase, EC 6.4.1.2) from three lines of Setaria viridis (L. Beauv.) resistant or sensitive to sethoxydim, and from one sethoxydim-sensitive line of Setaria italica (L. Beauv.) were cloned and sequenced. Sequence comparison revealed that a single isoleucine-leucine substitution discriminated ACCases from sensitive and resistant lines. Using near-isogenic lines of S. italica derived from interspecific hybridisation, we demonstrated that the transfer of the S. viridis mutant ACCase allele into a sethoxydim-sensitive S. italica line conferred resistance to this herbicide. We confirmed this result using allele-specific polymerase chain reaction and showed that a single copy of the mutant allele is sufficient to confer resistance to sethoxydim. We conclude that a mutant allele of chloroplastic ACCase encoding a leucine residue instead of an isoleucine residue at position 1780 is a major gene of resistance to sethoxydim.     

Genetic structure of landraces in foxtail millet (Setaria italica (L.) P. Beauv.) revealed with transposon display and interpretation to crop evolution of foxtail millet

Although the origin and domestication process of foxtail millet (Setaria italica subsp. italica (L.) P. Beauv.) has been studied by several groups, the issue is still ambiguous. It is essential to resolve this issue by studying a large number of accessions with sufficient markers covering the entire genome. Genetic structures were analyzed by transposon display (TD) using 425 accessions of foxtail millet and 12 of the wild ancestor green foxtail (Setaria italica subsp. viridis (L.) P. Beauv.). We used three recently active transposons (TSI-1, TSI-7, and TSI-10) as genome-wide markers and succeeded in demonstrating geographical structures of the foxtail millet. A neighbor-joining dendrogram based on TD grouped the foxtail millet accessions into eight major clusters, each of which consisted of accessions collected from adjacent geographical areas. Eleven out of 12 green foxtail accessions were grouped separately from the clusters of foxtail millet. These results indicated strong regional differentiations and a long history of cultivation in each region. Furthermore, we discuss the relationship between foxtail millet and green foxtail and suggest a monophyletic origin of foxtail millet domestication.     

谷子全生育期抗旱性鉴定及抗旱指标筛选

作物全生育期抗旱性对发掘和利用抗旱品种和抗旱基因至关重要,但谷子全生育期抗旱鉴定至今未有报道,抗旱鉴定缺乏鉴定指标。本试验在干旱池模拟干旱条件下,对苗期抗旱性表现不同的谷子品种进行了全生育期抗旱性研究,调查分析了根干重等形态和生理性状在干旱胁迫条件下的变化;采用相关和灰色关联度等方法,分析这些指标与抗旱性(DRI)的关系。结果表明,相对根冠比、相对单穗粒重和灌浆期光合速率、蒸腾速率同抗旱性表现极显著相关,可以作为谷子全生育期抗旱性鉴定的指标;而相对根干重、相对单穗重、相对株高和气孔导度则可以作为谷子全生育期抗旱性鉴定的参考指标;在供试的品种中,红根谷和大齐头白表现了良好的综合抗旱性。本文还讨论了抗旱性的复杂性以及不同性状对干旱胁迫的反应,所建立的指标对谷子全生育期抗旱鉴定具有指导意义。     

抗拿捕净晋谷21突变体的筛选和分子鉴定

谷子(Setaria italica(L.)P.Beauv.)是我国重要的杂粮作物,具有抗旱耐贫瘠、水分利用率高、适应性广、营养丰富等特点,但谷子田间除草一直制约着谷子的集约化栽培和有机旱作产业规模化的推广与发展。为快速选育抗除草剂谷子品种,本试验以晋谷21突变体M2群体为试验材料,通过喷施0.33%拿捕净除草剂对晋谷21突变群体进行初次筛选和再次筛选,进而快速获得抗拿捕净除草剂突变体植株并对其进行分子鉴定。结果显示,在大田苗期喷施0.33%拿捕净除草剂的初次筛选中,筛选出抗拿捕净除草剂的晋谷21突变体共39个株系;对这39个突变体株系再次喷施0.33%拿捕净除草剂,筛选获得9个有抗拿捕净除草剂特性的株系。对筛选获得的抗拿捕净突变体株系的其中3个株系共15个植株进行分子鉴定,发现有4个突变体植株的ACCase基因编码的第1780个氨基酸-异亮氨酸突变为亮氨酸。该结果可为今后利用分子生物学手段改良名优谷子品种晋谷21、加速抗除草剂谷子品种的选育提供理论和材料基础。     

The characterization of phenylalanine ammonia-lyase from several plant species

Inhibition of the enzyme phenylalanine ammonia-lyase is considered as a target for the design of herbicides. A reliable and simple assay for the enzyme has been used and the kinetics of the enzyme from several sources compared. Purification of the enzyme from the grass green foxtail (Setaria glauca) did not change its kinetic behavior. The distribution of phenylalanine ammonia-lyase and tyrosine ammonia-lyase activity in various plant species was determined.     

The molecular bases for resistance to acetyl co-enzyme A carboxylase (ACCase) inhibiting herbicides in two target-based resistant biotypes of annual ryegrass (Lolium rigidum)

Acetyl-CoA carboxylase (ACCase) (EC.6.4.1.2) is an essential enzyme in fatty acid biosynthesis and, in world agriculture, commercial herbicides target this enzyme in plant species. In nearly all grass species the plastidic ACCase is strongly inhibited by commercial ACCase inhibiting herbicides [aryloxyphenoxypropionate (APP) and cyclohexanedione (CHD) herbicide chemicals]. Many ACCase herbicide resistant biotypes (populations) of L. rigidum have evolved, especially in Australia. In many cases, resistance to ACCase inhibiting herbicides is due to a resistant ACCase enzyme. Two ACCase herbicide resistant L. rigidum biotypes were studied to identify the molecular basis of ACCase inhibiting herbicide resistance. The carboxyl-transferase (CT) domain of the plastidic ACCase gene was amplified by PCR and sequenced. Amino acid substitutions in the CT domain were identified by comparison of sequences from resistant and susceptible plants. The amino acid residues Gln-102 (CAG codon) and Ile-127 (ATA codon) were substituted with a Glu residue (GAG codon) and Leu residue (TTA codon), respectively, in both resistant biotypes. Amino acid positions 102 and 127 within the fragment sequenced from L. rigidum corresponded to amino acid residues 1756 and 1781, respectively, in the A. myosuroides full ACCase sequence. Allele-specific PCR results further confirmed the mutations linked with resistance in these populations. The Ile-to-Leu substitution at position 1781 has been identified in other resistant grass species as endowing resistance to APP and CHD herbicides. The Gln-to-Glu substitution at position 1756 has not previously been reported and its role in herbicide resistance remains to be established.     

SNP markers for black-grass ( Alopecurus myosuroides Huds.) genotypes resistant to acetyl CoA-carboxylase inhibiting herbicides

Chloroplastic acetyl CoA-carboxylase (ACCase) is the target of widely used, specific graminicide herbicides: cyclohexanediones (CHDs) and aryloxyphenoxypropionates (APPs). Resistance to these compounds is a worldwide, increasing problem. Population genetic studies aimed at understanding the dynamics of this situation and the diffusion of resistance genes within and between weed populations are challenging because biological assays are not adequate for this purpose, and because different mechanisms of resistance confer a similar resistance phenotype. Molecular markers for specifically detecting resistance genes are therefore urgently needed to conduct such studies. For this purpose, we cloned and sequenced the whole gene encoding chloroplastic ACCase in Alopecurus myosuroides Huds. (Black-grass). We identified two point mutations at nucleotide 5,341 that both cause an isoleucine-leucine substitution at position 1,781. Three bi-directional allele-specific PCR assays were developed, each detecting two distinct ACCase alleles with a single PCR reaction. The sensitivity of 1,190 seedlings of A. myosuroides to one CHD and one APP was determined. Genotyping revealed that, although resistant plants were only selected by APPs, the (1,781)Leu ACCase allele is a widespread, dominant gene of resistance to both APPs and CHDs. No other ACCase allele associated with resistance could be identified in this work. Useful applications of allele-specific PCR markers are population genetic studies as well as routine molecular diagnosis of herbicide resistance.     

Silicon Deposition in the Inflorescence Bristles and Macrohairs of Setaria italica (L.) Beauv.

Silicon deposits in the inflorescence bristles subtending eachspikelet, and in the macrohairs of foxtail millet (Setaria italica(L.) Beauv.) were investigated using scanning electron microscopyand electron-probe microanalysis. High concentrations of silicon were detected in the pricklehairs which covered the bristles. In the unicellular macrohairscovering the inflorescence axis and its branches, silicon wasdeposited along the whole length of the hairs. The mechanisms by which silicification may have taken place,and the possibility that the bristles and macrohairs are involvedin the aetiology of oesophageal cancer in N. China are discussed. Setaria italica (L.) Beauv., foxtail millet, millet, silicon deposits, silicification, prickle hairs, scanning electron microscopy, electron-probe microanalysis     

不同生态区主要育成谷子品种芽期耐旱性鉴定

以-0.5 MPa的PEG6000作渗透介质模拟干旱条件,对不同生态区的谷子(Setaria italica(L.)Beauv.)品种(系)进行种子萌发耐旱鉴定。结果表明:在PEG6000胁迫下,萌发耐旱指数与相对根芽比、芽生长抑制率呈极显著负相关;与根生长抑制率呈正相关,但相关系数小,相关不显著;与相对发芽势呈极显著正相关,且相关系数达0.939,可以反映谷子芽期耐旱性。根据萌发耐旱指数,将201份谷子品种(系)划分为极抗旱、抗旱、中度抗旱、不抗旱和极不抗旱5个等级。     

A Ser–Gly substitution in plastid-encoded photosystem II D1 protein is responsible for atrazine resistance in foxtail millet (<Emphasis Type="Italic">Setaria italica</Emphasis>)

A foxtail millet (Setaria italica L. Beauv.) line resistant to atrazine was obtained through interspecific hybridization between wild S. viridis L. Beauv. and cultivated S. italica. The resistance was proved to be controlled by a chloroplast-inherited gene and it has further been utilized in foxtail millet production. However, the sequence information of the putative atrazine resistance gene, psbA in foxtail millet’s chloroplast genome encoding photosystem II D1 protein (32 kDa thylakoid membrane protein) (photosystem Q_B protein) and the mutation site responsible for the resistance are not known. In this paper the psbA sequences of six atrazine susceptible/resistant foxtail millet varieties were obtained and compared. The results indicated that there was only one amino acid difference between susceptible and resistance gene, resulting from a single base substitution. It was concluded that a mutant allele of photosystem II protein D1 encoding a Gly residue instead of a Ser residue at position 264 is a major gene of resistance to atrazine. Moreover, the phylogenetic tree based on the psbA coding region of thirty-five plant species was carried out. The phylogenetic relationship between S. italica and other plants and the related evolutionary issues were discussed and it was suggested that psbA sequences could be used in phylogenetic studies in plants. Xiaoping Jia and Jincheng Yuan have equal contribution.     

禾谷类作物抗倒性研究方法与谷子抗倒性评价

倒伏是植株与其生长的环境条件相互作用的结果,是禾谷类作物产量和品种的重要限制因素。诱发倒伏的自然条件不是年年发生,但发生的程度差异很大,难以在自然条件下重复评价作物的倒伏性,因而人工诱发倒伏对于作物抗倒性的评价十分重要。谷子是非常容易发生倒伏的禾谷类作物,抗倒性的强弱是决定谷子高产、稳产的关键因素之一。由于谷子抗倒性的研究相对较少,本文通过对禾谷类作物抗倒伏性研究方法和评价指标的述评,为谷子抗倒性的研究提供借鉴,并对谷子倒伏与植株和产量性状的关系、品种抗倒性评价的指标等相关研究的进展做一综述。     

The resistance of plants Setaria veridis (L.) Beauv. to the influence of cadmium

It was shown that cadmium causes a slowdown of green foxtail Setaria veridis (L.) Beauv. shoots; however it had no effect on root growth or accumulation of underground and above-ground biomass. In the analysis of the effect of cadmium on the water regime and photosynthesis of plants, it was found that it had a negative effect on the stomatal apparatus, which led to a decrease in the transpiration rate and stomatal conductance. It was noted that the water content of tissues, as well as the rate of photosynthesis in the presence of cadmium, remained at the level of those in control plants. A high resistance of green foxtail to the effects of cadmium was established, which is provided with different adaptive mechanisms and anatomical and physiological characteristics associated with its affiliation to the group of C₄-plants.     

Silica Distribution in the Caryopsis and Inflorescence Bracts of Foxtail Millet [Setaria italica (L.) Beauv.] and its Possible Significance in Carcinogenesis

Silicon deposits in the caryopsis and inflorescence bracts offoxtail millet [Setaria italica (L.) Beauv.] were investigatedusing light microscopy, scanning electron microscopy and electronprobe microanalysis. The samples were obtained from Lin Xian,Henan province, northern China and CSIRO, Australia. High concentrations of silicon were observed in the papillaeon the external surfaces of the inflorescence bracts, and inthe epidermal cells. In the caryopsis silicon was depositedin the aleurone layer. Silicification was heavier in the LinXian samples. The heavy accumulation of silica in the foxtail millet and thepossibility that fragments of plant silica may be implicatedin the aetiology of oesophageal cancer in the Lin Xian regionare discussed. Setaria italica (L.), Beauv, foxtail millet, caryopsis, silica distribution, scanning electron microscopy     

Phytolith analysis for differentiating between foxtail millet (Setaria italica) and green foxtail (Setaria viridis)

Foxtail millet (Setaria italica) is one of the oldest domesticated cereal crops in Eurasia, but identifying foxtail millets, especially in charred grains, and differentiating it from its wild ancestor, green foxtail (Setaria viridis), in the archaeobotanical remains, is still problematic. Phytolithic analysis provides a meaningful method for identifying this important crop. In this paper, the silicon structure patterns in the glumes, lemmas, and paleas from inflorescence bracts in 16 modern plants of foxtail millet and green foxtail from China and Europe are examined using light microscopy with phase-contrast and a microscopic interferometer. Our research shows that the silicon structure of ΩIII from upper lemmas and paleas in foxtail millet and green foxtail can be correspondingly divided into two groups. The size of ΩIII type phytolith of foxtail millet is bigger than that from green foxtail. Discriminant function analysis reveals that 78.4% of data on foxtail millet and 76.9% of data on green foxtail are correctly classified. This means certain morphotypes of phytoliths are relatively reliable tools for distinguishing foxtail millet from green foxtail. Our results also revealed that the husk phytolith morphologies of foxtail millets from China and Eastern Europe are markedly different from those from Western Europe. Our research gives a meaningful method of separating foxtail millet and green foxtail. The implications of these findings for understanding the history of foxtail millet domestication and cultivation in ancient civilizations are significant.     

Colonization and bioherbicidal activity on green foxtail by Pseudomonas fluorescens BRG100 in a pesta formulation

Pseudomonas fluorescens BRG100 produces secondary metabolites with herbicidal activity on green foxtail ( Setaria viridis ), an important weed pest in Canadian agriculture. Five gfp transformants of P. fluorescens BRG100 were compared with the wild-type isolate for green foxtail root herbicide activity, i.e., root growth suppression, doubling time, carbon utilization, and colonization of green foxtail root (proximal and distal regions). The most revealing comparison between the wild type and its gfp transformants was herbicidal activity on green foxtail. Herbicidal activity of transformant gfp-7 was not significantly different from the uninoculated control, suggesting that insertion of the gfp gene may have interfered with a gene, or genes, vital to the bioherbicide process. Doubling time, carbon utilization, and colonization of green foxtail did not differ to a great extent between the wild type and the gfp transformants, indicating their suitability as conservatively tagged organisms for subsequent colonization-herbicidal activity studies. Accordingly, a pesta granule formulation delivered transformant gfp-2 to the seed coat and roots of green foxtail. Epifluorescent and confocal laser scanning microscopy revealed the transformant gfp-2 colonized the ventral portion of the seed coat, root hairs, and all areas of the root except the root cap region, where gfp-2 presumably exerted herbicidal effects. These results suggest that P. fluorescens BRG100 has considerable potential as a bioherbicide because of its successful colonization and suppressive activity on green foxtail root growth.     

Tubulin-isotype analysis of two grass species-resistant to dinitroaniline herbicides

Trifluralin-resistant biotypes of Eleusine indica (L.) Gaertn. (goosegrass) and Setaria viridis (L.) Beauv. (green foxtail) exhibit cross-resistance to other dinitroaniline herbicides. Since microtubules are considered the primary target site for dinitroaniline herbicides we investigated whether the differential sensitivity of resistant and susceptible biotypes of these species results from modified tubulin polypeptides. One-dimensional and two-dimensional polyacrylamide gel electrophoresis combined with immunoblotting using well-characterised anti-tubulin monoclonal antibodies were used to display the family of tubulin isotypes in each species. Seedlings of E. indica exhibited four -tubulin isotypes and one -tubulin isotype, whereas those of S. viridis exhibited two -tubulin and two -tubulin isotypes. Comparison of the susceptible and resistant biotypes within each species revealed no differences in electrophoretic properties of the multiple tubulin isotypes. These results provide no evidence that resistance to dinitroaniline herbicides is associated with a modified tubulin polypeptide in these biotypes of E. indica or S. viridis.Abbreviations 1-D, 2-D one-, two-dimensional - ID₅₀ dose causing 50% inhibition - PAGE polyacrylamide gel electrophoresis - SDS sodium dodecyl sulfate - S susceptible - R resistant We gratefully acknowledge Professor B.J. Gossett (Clemson University, South Carolina, USA) and Professor I.N. Morrison (University of Manitoba, Winnipeg, Canada) for providing seed stocks of Eleusine indica and Setaria viridis, respectively. Antibodies TAT-1 and KMX-1 were a kind gift from Professor K. Gull (University of Manchester, UK). Technical assistance with seed germination was provided by Ms. K.E. Cronin. We acknowledge financial support from the Science and Engineering Research Council for a studentship to T.R.W.     

The Ultrastructure and Analytical Microscopy of Silicon Deposition in the Aleurone Layer of the Caryopsis of Setaria italica (L.) Beauv.

Silicon deposition in the caryopsis of foxtail millet (Setariaitalica (L.) Beauv.) from Lin Xian, Northern China was investigatedusing transmission electron microscopy and energy dispersiveX-ray analysis. The highest silicon count rates were obtained from the pericarpand outer aleurone cell walls, and particularly from a granularelectron-opaque layer external to the outer aleurone cell wall.Silicon was not detected in tissues interior to the aleurone. A possible mechanism for silicon deposition in the caryopsisis suggested, and the results are discussed with respect tothe high incidence of oesophageal cancer in Lin Xian, NorthernChina. Setaria italica (L.) Beauv., foxtail millet, silicon deposition, alcurone layer, caryopsis, ultrastructure, X-ray analysis     

谷子叶绿体乙酰辅酶A羧化酶cDNA的克隆和禾本科除草剂靶位点的序列分析

乙酰辅酶A羧化酶是一个生物素羧化酶,它所催化的反应是脂肪酸生物合成中的第一个植物叶绿体中的乙酰辅酶A羧化酶是两类禾本科除草剂的靶蛋白.从抗除草剂拿捕净和感拿捕净的谷子(SetariaitalicaBeauv.)中克隆了两个乙酰辅酶A羧化酶的全长cDNA,分别命名为foxACC-R和foxACC-S,它们推导的蛋白质均编码2 321个氨基酸,然而在第1 780个氨基酸处,foxACC-R编码亮氨酸,而foxACC-S编码异亮氨酸.采用生物信息学方法,我们推断这个cDNA编码的是叶绿体中的乙酰辅酶A羧化酶,并预测了它的功能域和保守区.通过这两个cDNA编码的氨基酸序列与其他乙酰辅酶A羧化酶的序列比较得出结论,亮氨酸/异亮氨酸位点可能是APPs和CHDs两类除草剂作用的关键位点.Southern杂交分析的结果显示,该基因在谷子基因组中只有一个拷贝.