Evolution of Ac and Dsl elements in select grasses (Poaceae)

We present data on evolution of the Ac/Ds family of transposable elements in select grasses (Poaceae). An Ac-like element was cloned from a DNA library of the grass Pennisetum glaucum (pearl millet) and 2387 bp of it have been sequenced. When the pearl millet Ac-like sequence is aligned with the corresponding region of the maize Ac sequence, it is found that all sequences corresponding to intron II in maize Ac are absent in pearl millet Ac. Kimura's evolutionary distance between maize and pearl millet Ac sequences is estimated to be 0.429±0.020 nucleotide substitutions per site. This value is not significantly different from the average number of synonymous substitutions for coding regions of the Adh1 gene between maize and pearl millet, which is 0.395±0.051 nucleotide substitutions per site. If we can assume Ac and Adh1 divergence times are equivalent between maize and pearl millet, then the above calculations suggest Ac-like sequences have probably not been strongly constrained by natural selection. The level of DNA sequence divergence between maize and pearl millet Ac sequences, the estimated date when maize and pearl millet diverged (25–40 million years ago), coupled with their reproductive isolation/lack of current genetic exchange, all support the theory that Ac-like sequences have not been recently introduced into pearl millet from maize. Instead, Ac-like sequences were probably present in the progenitor of maize and pearl millet, and have thus existed in the grasses for at least 25 million years. Ac-like sequences may be widely distributed among the grasses. We also present the first 2 Dsl controlling element sequences from teosinte species: Zea luxurians and Zea perennis. A total of 10 Dsl elements had previously been sequenced from maize and a distant maize relative, Tripsacum. When a maximum likelihood network of genetic relationships is constructed for all 12 sequenced Dsl elements, the 2 teosinte Dsl elements are as distant from most maize Dsl elements and from each other, as the maize Dsl elements are from one another. Our new teosinte sequence data support the previous conclusion that Dsl elements have been accumulating mutations independently since maize and Tripsacum diverged. We present a scenario for the origin of Dsl elements.     

Molecular evolutionary characterization of an Activator (Ac)-like transposable element sequence from pearl millet (Pennisetum glaucum) (Poaceae)

We present data on the evolution of the Ac/Ds family of transposable elements in select grasses (Poaceae). A defective Ac-like element was cloned from a DNA library of the grass Pennisetum glaucum (pearl millet) and its entire 4531 bp sequence has been determined. When the pearl millet Ac-like sequence is aligned with the maize Ac sequence, it is found that there is approximately 70% DNA similarity in the central region spanning most of maize Ac exon II and all of exon III. In addition, there are two smaller regions of similarity at the Ac terminii. Besides these three major structural similarities, Pennisetum Ac has two large regions, one 5 and one 3, that show little similarity to Zea Ac. Furthermore, most of the sequences corresponding to intron II in maize Ac are absent in pearl millet Ac. Kimura's evolutionary distance between the central region of maize and pearl millet Ac sequences is estimated to be 0.429±0.020 nucleotide substitutions per site. This value is not significantly different from the average number of synonymous substitutions for coding regions of the Adh1 gene between maize and pearl millet, which is 0.395±0.051 nucleotide substitutions per site. If we assume Ac and Adh1 divergence times are equivalent between maize and pearl millet, then the above calculations suggest Ac-like sequences have probably not been strongly constrained by natural selection. Conserved DNA and amino acid sequence motifs are also examined. The level of DNA sequence divergence between maize and pearl millet Ac sequences, the estimated date when maize and pearl millet diverged (25–40 million years ago), coupled with their reproductive isolation/lack of current genetic exchange, all support the theory that Ac-like sequences have not been recently introduced into pearl millet from maize. Instead, Ac-like sequences were probably present in the progenitor of maize and pearl millet and have thus existed in the grasses for at least 25 million years.     

Polymorphism pattern at a miniature inverted‐repeat transposable element locus downstream of the domestication gene Teosinte‐branched1 in wild and domesticated pearl millet

Unravelling the mechanisms involved in adaptation to understand plant morphological evolution is a challenging goal. For crop species, identification of molecular causal polymorphisms involved in domestication traits is central to this issue. Pearl millet, a domesticated grass mostly found in semi‐arid areas of Africa and India, is an interesting model to address this topic: the domesticated form shares common derived phenotypes with some other cereals such as a decreased ability to develop basal and axillary branches in comparison with the wild phenotype. Two recent studies have shown that the orthologue of the maize gene Teosinte‐Branched1 in pearl millet (PgTb1) was probably involved in branching evolution during domestication and that a miniature inverted‐repeat transposable element (MITE) of the Tuareg family was inserted in the 3′ untranslated region of PgTb1. For a set of 35 wild and domesticated populations, we compared the polymorphism patterns at this MITE and at microsatellite loci. The Tuareg insertion was nearly absent in the wild populations, whereas a strong longitudinal frequency cline was observed in the domesticated populations. The geographical pattern revealed by neutral microsatellite loci clearly demonstrated that isolation by distance does not account for the existence of this cline. However, comparison of population differentiation at the microsatellite and the MITE loci and analyses of the nucleotide polymorphism pattern in the downstream region of PgTb1 did not show evidence that the cline at the MITE locus has been shaped by selection, suggesting the implication of a neutral process. Alternative hypotheses are discussed.     

<Emphasis Type="Italic">hATpin</Emphasis>, a Family of MITE-like <Emphasis Type="Italic">hAT</Emphasis> Mobile Elements Conserved in Diverse Plant Species that Forms Highly Stable Secondary Structures

We identified a 178 bp mobile DNA element in lettuce with characteristic CGAGC/GCTCG repeats in the subterminal regions. This element has terminal inverted repeats and 8-bp target site duplications typical of the hAT superfamily of class II mobile elements, but its small size and potential to form a single-stranded stable hairpin-like secondary structure suggest that it is related to MITE elements. In silico searches for related elements identified 252 plant sequences with 8-bp target site duplications and sequence similarity in their terminal and subterminal regions. Some of these sequences were predicted to encode transposases and may be autonomous elements; these constituted a separate clade within the phylogram of hAT transposases. We demonstrate that the CGAGC/GCTCG pentamer maximizes the hairpin stability compared to any other pentamer with the same C + G content, and the secondary structures of these elements are more stable than for most MITEs. We named these elements collectively as hATpin elements because of the hAT similarity and their hairpin structures. The nearly complete rice genome sequence and the highly advanced genome annotation allowed us to localize most rice elements and to deduce insertion preferences. hATpin elements are distributed on all chromosomes, but with significant bias for chromosomes 1 and 10 and in regions of moderate gene density. This family of class II mobile elements is found primarily in monocot species, but is also present in dicot species. Electronic supplementary material Electronic supplementary material is available for this article at and accessible for authorised users.     

Recent amplification of miniature inverted-repeat transposable elements in the vector mosquito Culex pipiens: characterization of the Mimo family

We describe a new family of repetitive elements, named Mimo, from the mosquito Culex pipiens. Structural characteristics of these elements fit well with those of miniature inverted-repeat transposable elements (MITEs), which are ubiquitous and highly abundant in plant genomes. The occurrence of Mimo in C. pipiens provides new evidence that MITEs are not restricted to plant genomes, but may be widespread in arthropods as well. The copy number of Mimo elements in C. pipiens (1000 copies in a 540 Mb genome) supports the hypothesis that there is a positive correlation between genome size and the magnitude of MITE proliferation. In contrast to most MITE families described so far, members of the Mimo family share a high sequence conservation, which may reflect a recent amplification history in this species. In addition, we found that Mimo elements are a frequent nest for other MITE-like elements, suggesting that multiple and successive MITE transposition events have occurred very recently in the C. pipiens genome. Despite evidence for recent mobility of these MITEs, no element has been found to encode a protein; therefore, we do not know how they have transposed and have spread in the genome. However, some sequence similarities in terminal inverted-repeats suggest a possible filiation of some of these mosquito MITEs with pogo-like DNA transposons.     

Conservation and Diversification of <Emphasis Type="Italic">SCARECROW</Emphasis> in Maize

The SCARECROW (SCR) gene in Arabidopsis is required for asymmetric cell divisions responsible for ground tissue formation in the root and shoot. Previously, we reported that Zea mays SCARECROW (ZmSCR) is the likely maize ortholog of SCR. Here we describe conserved and divergent aspects of ZmSCR. Its ability to complement the Arabidopsis scr mutant phenotype suggests conservation of function, yet its expression pattern during embryogenesis and in the shoot system indicates divergence. ZmSCR expression was detected early during embryogenesis and localized to the endodermal lineage in the root, showing a gradual regionalization of expression. Expression of ZmSCR appeared to be analogous to that of SCR during leaf formation. However, its absence from the maize shoot meristem and its early expression pattern during embryogenesis suggest a diversification of ZmSCR in the patterning processes in maize. To further investigate the evolutionary relationship of SCR and ZmSCR, we performed a phylogenetic analysis using Arabidopsis, rice and maize SCARECROW-LIKE genes (SCLs). We found SCL23 to be the most closely related to SCR in both eudicots and monocots, suggesting that a gene duplication resulting in SCR and SCL23 predates the divergence of dicots and monocots. Electronic supplementary material Electronic supplementary material is available for this article at and accessible for authorised users.     

Regulation of the <Emphasis Type="Italic">cellulose synthase-like</Emphasis> gene family by light in the maize mesocotyl

The cellulose synthase-like (ZmCSL) gene family of maize was annotated and its expression studied in the maize mesocotyl. A total of 28 full-length CSL genes and another 13 partial sequences were annotated; four are predicted to be pseudogenes. Maize has all of the CSL subfamilies that are present in rice, but the CSLC subfamily is expanded from 6 in rice to 12 in maize, and the CSLH subfamily might be reduced from 3 to 1. Unlike rice, maize has a gene in the CSLG subfamily, based on its sequence similarity to two genes annotated as CSLG in poplar. Light regulation of glycan synthase enzyme activities and CSL gene expression were analyzed in the mesocotyl. A Golgi-localized glucan synthase activity is reduced by ~50% 12 h after exposure to light. β-1,4-Mannan synthase activity is reduced even more strongly (>85%), whereas β-1,4-xylan synthase, callose synthase, and latent IDPase activity respond only slightly, if at all, to light. At least 17 of the CSL genes (42%) are expressed in the mesocotyl, of which four are up-regulated at least twofold, seven are down-regulated at least twofold, and six are not affected by light. The results contribute to our understanding of the structure of the CSL gene family in an important food and biofuel plant, show that a large percentage of the CSL genes are expressed in the specialized tissues of the mesocotyl, and demonstrate that members of the CSL gene family are differentially subject to photobiological regulation. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Pearl millet transformation system using the positive selectable marker gene phosphomannose isomerase

Fertile transgenic pearl millet plants expressing a phosphomannose isomerase (PMI) transgene under control of the maize ubiquitin constitutive promoter were obtained using the transformation system described here. Proliferating immature zygotic embryos were used as target tissue for bombardment using a particle inflow gun. Different culture and selection strategies were assessed in order to obtain an optimised mannose selection protocol. Stable integration of the manA gene into the genome of pearl millet was confirmed by PCR and Southern blot analysis. Stable integration of the manA transgene into the genome of pearl millet was demonstrated in T₁ and T₂ progeny of two independent transformation events with no more than four to ten copies of the transgene. Similar to results obtained from previous studies with maize and wheat, the manA gene was shown to be a superior selectable marker gene for improving transformation efficiencies when compared to antibiotic or herbicide selectable marker genes.Abbreviations 2,4-D: 2,4-Diclorophenoxyacetic acid - IAA: Indole acetic acid - ICRISAT: International Crops Research Institute for the Semi-Arid Tropics - IZEs Immature zygotic embryos Communicated by H. Lörz     

<Emphasis Type="Italic">Mutator</Emphasis>-like elements identified in melon,Arabidopsis and rice contain ULP1 protease domains

The transposon Mutator was first identified in maize, and is one of the most active mobile elements in plants. The Arabidopsis thaliana genome contains at least 200 Mutator-like elements (MULEs), which contain the Mutator-like transposase gene, and often additional genes. We have detected a novel type of MULEs in melon (CUMULE), which, besides the transposase, contains two ubiquitin-like specific protease-like sequences (ULP1). This element is not present in the observed location in some melon cultivars. Multiple copies of this element exist in the Cucumis melo genome, and it has been detected in other Cucurbitaceae species. Analysis of the A. thaliana genome revealed more than 90 CUMULE-like elements, containing one or two Ulp1-like sequences, although no evidence of mobility exists for these elements. We detected various putative transposable elements containing ULP1-like sequences in rice. The discovery of these MULEs in melon and Arabidopsis, and the existence of similar elements in rice and maize, suggest that a proteolytic function may be important for this subset of the MULE transposable elements. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users. Nucleotide sequence data reported are available in the GenBank database under the accession number AY524004.     

<Emphasis Type="Italic">Drosophila</Emphasis><Emphasis Type="Italic">P</Emphasis> transposons of the urochordata <Emphasis Type="Italic">Ciona intestinalis</Emphasis>

P transposons belong to the eukaryotic DNA transposons, which are transposed by a cut and paste mechanism using a P-element-coded transposase. They have been detected in Drosophila, and reside as single copies and stable homologous sequences in many vertebrate species. We present the P elements Pcin1, Pcin2 and Pcin3 from Ciona intestinalis, a species of the most primitive chordates, and compare them with those from Ciona savignyi. They showed typical DNA transposon structures, namely terminal inverted repeats and target site duplications. The coding region of Pcin1 consisted of 13 small exons that could be translated into a P-transposon-homologous protein. C. intestinalis and C. savignyi displayed nearly the same phenotype. However, their P elements were highly divergent and the assumed P transposase from C. intestinalis was more closely related to the transposase from Drosophila melanogaster than to the transposase of C. savignyi. The present study showed that P elements with typical features of transposable DNA elements may be found already at the base of the chordate lineage. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Expressional profiling study revealed unique expressional patterns and dramatic expressional divergence of maize α-zein super gene family

The α-zein super gene family encodes the most predominant storage protein in maize (Zea mays) endosperm. In maize inbred line B73, it consists of four gene families with 41 member genes. In this study, we combined quantitative real-time PCR and random clone sequencing to successfully profile the expression of α-zein super gene family during endosperm development. We found that only 18 of the 41 member genes were expressed, and their expression levels diverge greatly. At the gene family level, all families had characteristic “up-and-down” oscillating expressional patterns that diverged into two major groups. At the individual gene level, member genes showed dramatic divergence of expression patterns, indicating fast differentiation of their expression regulation. A comparison study among different inbred lines revealed significantly different expressed gene sets, indicating the existence of highly diverged haplotypes. Large gene families containing long gene clusters, e.g. z1A or z1C, mainly contributed the highly divergent haplotypes. In addition, allelic genes also showed significant divergence in their expressional levels. These results indicated a highly dynamic and fast evolving nature to the maize α-zein super gene family, which might be a common feature for other large gene families. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Lingna Feng and Jia Zhu contributed equally to this work.     

谷子PAL基因家族全基因组的鉴定和表达分析

苯丙氨酸解氢酶(phenylalanine ammonia-lyase,PAL)基因家族参与苯丙烷类代谢过程,通过调控植物抗病次生物质的合成在植物抗逆反应中发挥重要作用。为明确谷子PAL基因家族在逆境胁迫下的表达规律,该研究利用生物信息学方法对谷子PAL基因家族进行鉴定和表达分析。结果表明:谷子具有11个PAL基因,在进化树中可分为3个亚家族,SiPAL7独自进化为一支。通过构建蛋白结构域发现PAL基因家族成员均含有保守的PAL结构域。启动子分析显示,PAL基因含有应答激素、逆境胁迫等多种因子的顺式作用元件,说明PAL基因广泛参与不同生物学调控过程。RT-qPCR结果显示,谷子PAL基因家族多为诱导型表达,不同光照条件下PAL基因表达量变化明显,不同基因具有不同响应模式,说明谷子PAL基因家族在参与光调节反应中发挥重要作用。谷子PAL基因高度保守,广泛响应不同非生物胁迫,具有表达特异性。该研究结果为揭示PAL基因家族在调节谷子抗性及胁迫应答过程中的作用提供了参考。     

Allelic variation of the Waxy gene in foxtail millet [Setaria italica (L.) P. Beauv.] by single nucleotide polymorphisms

The Waxy (Wx) gene product controls the formation of a straight chain polymer of amylose in the starch pathway. Dominance/recessiveness of the Wx allele is associated with amylose content, leading to non-waxy/waxy phenotypes. For a total of 113 foxtail millet accessions, agronomic traits and the molecular differences of the Wx gene were surveyed to evaluate genetic diversities. Molecular types were associated with phenotypes determined by four specific primer sets (non-waxy, Type I; low amylose, Type VI; waxy, Type IV or V). Additionally, the insertion of transposable element in waxy was confirmed by ex1/TSI2R, TSI2F/ex2, ex2int2/TSI7R and TSI7F/ex4r. Seventeen single nucleotide polymorphims (SNPs) were observed from non-coding regions, while three SNPs from coding regions were non-synonymous. Interestingly, the phenotype of No. 88 was still non-waxy, although seven nucleotides (AATTGGT) insertion at 2,993 bp led to 78 amino acids shorter. The rapid decline of r ² in the sequenced region (exon 1–intron 1–exon 2) suggested a low level of linkage disequilibrium and limited haplotype structure. K _s values and estimation of evolutionary events indicate early divergence of S. italica among cereal crops. This study suggested the Wx gene was one of the targets in the selection process during domestication. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Expression and functional analysis of <Emphasis Type="Italic">ZmDWF4</Emphasis>, an ortholog of <Emphasis Type="Italic">Arabidopsis DWF4</Emphasis> from maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.)

DWF4 encodes a rate-limiting mono-oxygenase that mediates 22α-hydroxylation reactions in the BR biosynthetic pathway and it is the target gene in the BR feedback loop. Knockout of DWF4 results in a dwarfed phenotype and other severe defects in Arabidopsis. Here we report on the isolation of the ZmDWF4 gene in maize. Sequence analysis revealed that the open reading frame of ZmDWF4 was 1,518 bp, which encodes a protein composed of 505 amino acid residues with a calculated molecular mass of 57.6 kD and a predicated isoelectric point (pI) of 9.54. Phylogenetic analysis indicated that ZmDWF4 was very close to the Arabidopsis DWF4. In young maize seedlings, the expression of ZmDWF4 in shoots was much higher than that in roots. The highest expression of ZmDWF4 was observed in husk leaves and the lowest in silks during flowering stage. The expression of ZmDWF4 in maize was significantly down regulated by exogenous brassinolide. A heterogeneous complementary experiment demonstrated that the defects of three Arabidopsis DWF4 mutants could be rescued by constitutive expression of ZmDWF4, with leaf expandability, inflorescence stem heights and fertile capabilities all restored to normal levels. Increases in seed and branch number as well as the height of florescence stem were observed in the over-expressed transformants. These findings suggest that ZmDWF4 may be an ortholog gene of Arabidopsis DWF4 and responsible for BR biosynthesis in maize. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Multiple subfamilies of mariner transposable elements are present in stalk-eyed flies (Diptera: Diopsidae)

The Diopsid stalk-eyed flies are an increasingly well-studied group. Presented here is evidence of the first known transposable elements discovered in these flies. The vertumnana mariner subfamily was identified in the Diopsini tribe, but could not be amplified in species of the Sphyracephalini tribe. PCR screening with degenerate primers revealed that multiple mariner subfamilies are present within the Diopsidae. Most of the sequenced elements appear to be pseudogenes; however two subfamilies are shown to be evolving under purifying selection, raising the possibility that mariner is active in some Diopsid species. Evidence is presented of a possible horizontal transfer event involving an unknown Teleopsis species and the Tephritid fly Bactrocera neohumeralis. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Molecular Evolution of the Ac/Ds Transposable-Element Family in Pearl Millet and Other Grasses

We report an Ac-like sequence from pearl millet (Pennisetum glaucum) and deletion derivative Ac-like sequences from pearl millet and another grass species, Bambusa multiplex. Sequence relationships between the pearl millet and maize Ac elements suggest the Ac/Ds transposable-element family is ancient. Further, the sequence identity between the Bambusa Ac-like sequence and maize Ac implies that the Ac/Ds transposable-element family has been in the grass family since its inception. The Ac-like sequences reported from pearl millet and maize Ac are statistically heterogeneous in pair-wise distance comparisons to each other. Yet, we are unable to discriminate between differential selection or ectopic exchange (recombination and conversion) between nonidentical transposable element homologues, as the cause of the heterogeneity. However, the more extreme heterogeneity exhibited between the previously described pearl millet element and maize Ac seems likely to derive from ectopic exchange between elements with different levels of divergence.     

Daily changes in nitrate influx,efflux and metabolism in maize and pearl millet

Maize (Zea mays L.) and pearl millet (Pennisetum americanum (L.) Leeke) seedlings were exposed to [¹⁵N]nitrate for 1-h periods at eight times during a 24-h period (16–8 h light-dark for maize; 14–10 h for millet). Influx of [¹⁵N]nitrate as well as its reduction and translocation were determined during each period. The efflux of previously absorbed [¹⁴N]nitrate to the uptake solution was also estimated. No marked diurnal changes in [¹⁴N]nitrate efflux or [¹⁵N]nitrate influx were evident in maize. In contrast, [¹⁴N]nitrate efflux from millet increased and eventually exceeded [¹⁵N]nitrate influx during the late dark and early light periods, resulting in net nitrate efflux from the roots. The dissimilarity of their diurnal patterns indicates that influx and efflux are independently regulated. In both species, [¹⁵N]nitrate reduction and ¹⁵N translocation to shoots were curtailed more by darkness than was [¹⁵N]nitrate influx. In the light, maize reduced 15% and millet 24% of the incoming [¹⁵N]nitrate. In darkness, reduction dropped to 11 and 17%, respectively. Since the accumulation of reduced-¹⁵N in shoots declined abruptly in darkness, whereas that in roots was little affected, it is suggested that in darkness [¹⁵N]nitrate reduction occurred primarily in roots. The decrease in nitrate uptake and reduction in darkness was not related to efflux, which remained constant in maize and did not respond immediately to darkness in pearl millet.Paper No. 6722 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh     

Overexpression of <Emphasis Type="Italic">Zm401</Emphasis>, an mRNA-like RNA,has distinct effects on pollen development in maize

We previously identified a 0.7 Kb cDNA fragment of Zm401, a novel pollen-specific gene in maize (Zea mays). However, little information is known about the function of Zm401 in pollen development. The full-length of Zm401 cDNA was amplified by 5′ RACE and 3′ RACE and both sequence analysis and in vitro translation of Zm401 showed that it belonged to an mRNA-like non-coding gene. To analyze its possible biological roles in pollen development, the Zm401 cDNA was overexpressed in transgenic maize under the control of a pollen specific promoter Zm13 or a CaMV 35S promoter. RT-PCR and RNA gel blot analysis indicated that the expression level of Zm401 in leaves and anthers of transgenic plants was much higher than that of non-transformants. Compared with the non-transformed maize, transgenic maize showed distinct phenotypes, such as abnormal tassels and degenerate anthers. The histological observation showed that the development of pollen grains and anthers in transgenic plants were abnormal. These abnormalities include delayed degradation of tapetum, asynchronous fusion of pollen sacs, and aborted pollen grain development. Furthermore, the pollen viability in six transgenic plants ranged from 1.24% to 6.63%. The reduced pollen viability cosegregated with the transgene in a selfed progeny. These results suggest that Zm401 is involved in the regulation of pollen development. This article demonstrated Zm401, as a non-coding RNA, plays an essential role in pollen development. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

A one-time inducible transposon for creating knockout mutants

The maize transposon Ac can move to a new location within the genome to create knockout mutants in transgenic plants. In rice, Ac transposon is very active but sometimes undergoes further transposition and leaves an empty mutated gene. Therefore, we developed a one-time transposon system by locating one end of the transposon in the intron of the Ac transposase gene, which is under the control of the inducible promoter (PR-1a). Treatment with salicylic acid induced transposition of this transposon, COYA, leading to transposase gene breakage in exons. The progeny plants inheriting the transposition events become stable knockout mutants, because no functional transposase could be yielded. The behavior of COYA was analyzed in single-copy transgenic rice plants. We determined the expression of the modified transposase gene and its ability to trigger transposition events in transgenic rice plants. The COYA element thus exhibits potential for development of an inducible transposon system suitable for gene isolation in heterologous plant species. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.