水稻双子房突变体中类copia逆转座子同源序列的研究

以水稻双子房突变体的总ＤＮＡ为模板,用简并引物扩增类ｃｏｐｉａ逆转座子的逆转录酶区域。从ＰＣＲ产物中分离到代表３个不同的类ｃｏｐｉａ逆转座子的片段,其中两个在水稻（Ｏｒｙｚａ ｓａｔｉｖａ Ｌ．）品种“窄叶青８号”和“京系１７”间产生的多态性杂交带分别定位于水稻７条染色体的９个位点。Ｒ３３－８含大量的终止码,Ｒ３３－１及Ｒ３３－４为连续的编码区,推测的氨基酸序列含８１个氨基酸残基。Ｒ３３－１的拷贝     

Transposable elements and the evolution of genome organization in mammals

All mammalian transposable elements characterized to date appear to be nonrandomly distributed in the mammalian genome. While no element has been found to be exclusively restricted in its chromosomal location, LINE elements and some retrovirus-like elements are preferentially accumulated in G-banding regions of the chromosomes, and in some cases in the sex chromosomes, while SINE elements occur preferentially in R-banding regions. Four mechanisms are presented which may explain the nonrandom genomic distribution of mammalian transposons: i) sequence-specific insertion, ii) S-phase insertion, iii) ectopic excision, and iv) recombinational editing. Some of the available data are consistent with each of these four models, but no single model is sufficient to explain all of the existing data.     

Transpositional landscape of the rice genome revealed by paired-end mapping of high-throughput re-sequencing data

Transposable elements (TEs) are mobile entities that densely populate most eukaryotic genomes and contribute to both their structural and functional dynamics. However, most TE-related sequences in both plant and animal genomes correspond to inactive, degenerated elements, due to the combined effect of silencing pathways and elimination through deletions. One of the major difficulties in fully characterizing the molecular basis of genetic diversity of a given species lies in establishing its genome-wide transpositional activity. Here, we provide an extensive survey of the transpositional landscape of a plant genome using a deep sequencing strategy. This was achieved through paired-end mapping of a fourfold coverage of the genome of rice mutant line derived from an in vitro callus culture using Illumina technology. Our study shows that at least 13 TE families are active in this genotype, causing 34 new insertions. This next-generation sequencing-based strategy provides new opportunities to quantify the impact of TEs on the genome dynamics of the species.     

Somaclonal variation in rice – Submergence tolerance and other agronomic characters

The ability to withstand complete submergence was studied in R₂ seedlings raised by self-pollination from 158 R₁ plants regenerated from callus of mature rice seeds (cultivars FR13A and Calrose). Compared to parental controls, significant improvement in submergence tolerance as assessed by percentage survival, number of new leaves and chlorophyll content of the third leaf, was found in 5 of the 115 cv. FR13A somaclones and 3 of the 43 cv. Calrose somaclones tested using an aquarium system in a controlled-temperature glasshouse. With some exceptions these improvements were also observed in the R₃ generation when tested under field conditions in Thailand. Variation in other agronomically important characters was observed in the R₂ plants grown in a temperature glasshouse under 2 contrasting environments.     

Germinal virus vector WDV (wheat dwarf virus)-mediated multiple insertions of a maize transposon,Ds (dissociation), in rice

Wheat dwarf virus (WDV) is a monocot-infecting geminivirus that replicates in infected tissue as double-stranded DIMA. We evaluated whether the WDV vector system bearingDs could be used as an effective insertional mutagen in rice. Molecular data showed thatDs was excised from WDV vectors once the WDV-carryingDs (WDV::Ds) and the genomicAc vector were co-introduced into rice calli. Mature TO and T1 transgenic plants were analyzed for the distribution and inheritance ofDs inserts. Southern analysis indicated that theDs elements excised from WDV vectors were stably inserted into genomes. The number of transposedDs ranged from zero to three copies, among independent transformants. Meanwhile, untransposedDs (WDV::Ds) were present in multiple-copies in genomes. Southern analysis of the selfed progeny of T0 plants demonstrated that most WDV::Ds were co-segregated among siblings. This indicated that these elements were integrated into the same single loci. However, a fewDs were found to segregate independently from the majority ofDs. In this report, we discuss the efficiency of WDV vectors in generating multicopyDs in rice genomes.     

Transposon dynamics and the breeding system

The selfish DNA hypothesis predicts that natural selection is responsible for preventing the unregulated build up of transposable elements in organismal genomes. Accordingly, between-species differences in the strength and effectiveness of selection against transposons should be important in driving the evolution of transposon activity and abundance. We used a modeling approach to investigate how the rate of self-fertilization influences the population dynamics of transposable elements. Contrasting effects of the breeding system were observed under selection based on transposon disruption of gene function versus selection based on element-mediated ectopic exchange. This suggests that the comparison of TE copy number in organisms with different breeding systems may provide a test of the relative importance of these forces in regulating transposon multiplication. The effects of breeding system also interacted with population size, particularly when there was no element excision. The strength and effectiveness of selection against transposons was reflected not only in their equilibrium abundance, but also in the per-site element frequency of individual insertions and the coefficient of variation in copy number. These results are discussed in relation to evidence on transposon abundance available from the literature, and suggestions for future data collection. With their immense variety of breeding systems,plants will be extremely important for comparative studies and for sorting out the forces influencing...variation. This revised version was published online in July 2006 with corrections to the Cover Date.     

Characterization of Some Chromosomal Aberrations in Regenerated Rice Plants (<Emphasis Type="Italic">Oryza sativa</Emphasis>)

Somaclonal variation was studied in two Iranian land races of O. sativa spp. japonica var Hassani and O. sativa spp. indica var Sadridomsiah and 2000 plants of each cultivar were cytogenetically examined in two steps. In the first step, chromosome counts of root tips was used to detect ploidy levels and aneuploids of regenerated plants. In the second step, chromosomal aberrations were characterized by pachytene analysis of PMC’s. Ploidy levels were seen between n and 4n (haploids to tetraploids) in both cultivars with diploid resource (2n = 2x = 24). The total rate of variation for Hassani (japonica) was 13.7% including 10.8% for changes in chromosome number (ploidy levels and aneuploids) and 2.9% for chromosomal aberrations such as deficiency. A total rate of variation for Sadridomsiah (indica) was 15.6% including 12.4% for change in numbers and 3.2% for aberrations in construction. Most of important cytological mutations were observed in various chromosomes among regenerated plants of cultivars. Neither nullisomics nor inversions were distinguished in any samples.     

Early and multiple Ac transpositions in rice suitable for efficient insertional mutagenesis

A GFP excision assay was developed to monitor the excision of Ac introduced into rice by Agrobacterium-mediated transformation. The presence of a strong double enhancer element of the CaMV 35S promoter adjacent to the Ac promoter induced very early excision, directly after transformation into the plant cell, exemplified by the absence of Ac in the T-DNA loci. Excision fingerprint analysis and characterization of transposition events from related regenerants revealed an inverse correlation between the number of excision events and transposed Ac copies, with single early excisions after transformation generating Ac amplification. New transpositions were generated at a frequency of 15–50% in different lines, yielding genotypes bearing multiple insertions, many of which were inherited in the progeny. The sequence of DNA flanking Ac in three representative lines provided a database of insertion tagged sites suitable for the identification of mutants of sequenced genes that can be examined for phenotypes in a reverse genetics strategy to elucidate gene function. Remarkably, two-thirds of Ac tagged sites showing homology to sequences in public databases were in predicted genes. A clear preference of transposon insertions in genes that are either predicted by protein coding capacity or by similarity to ESTs suggests that the efficiency of recovering knockout mutants of genes could be about three times higher than random. Linked Ac transposition, suitable for targeted tagging, was documented by segregation analysis of a crippled Ac element and by recovery of a set of six insertions in a contiguous sequence of 70 kb from chromosome 6 of rice.     

Mechanisms and rates of genome expansion and contraction in flowering plants

Plant genomes are exceptional for their great variation in genome size, an outcome derived primarily from their frequent polyploid origins and from the amplification of retrotransposons. Although most studies of plant genome size variation have focused on developmental or physiological effects of nuclear DNA content that might influence plant fitness, more recent studies have begun to investigate possible mechanisms for plant genome expansion and contraction. Analyses of relatively neutral genome components, like transposable elements, have been particularly fruitful, largely due to the enormous growth in genomic sequence information from many different plant species. Current data suggest that unequal recombination can slow the growth in genome size caused by retrotransposon amplification, but that illegitimate recombination and other deletion processes may be primarily responsible for the removal of non-essential DNA from small genome plants.     

RAPD analysis of blast resistant somaclones from upland rice cultivar IAC 47 for genetic divergence

Seventeen somaclones of upland rice cultivar IAC 47 showing different plant types, and either resistance or susceptibility to leaf blast, were utilized for random amplified polymorphic DNA (RAPD) analysis. Somaclones exhibited differences in reaction to isolates of Pyricularia grisea. Two somaclones (SC02 and SC04) were resistant to all three field isolates of somaclones, while the cultivar IAC 47 was susceptible. The inheritance study of two distinct plant types, one with erect bright green leaves and the other with droopy yellow green leaves, showed that a single possibly different, dominant gene governs each plant type. Of 32 random decamer primers utilized, OPA02 and OPD02 detected polymorphisms between somaclones showing erect bright green leaves and droopy yellow green leaves. Reliable grouping exhibiting 80% similarity was achieved with 17 primers. Leaf blast resistance to race IC-2 of P. grisea was associated with the plant type of erect bright green leaves.     

Evidence for genomic changes in transgenic rice (Oryza sativa L.) recovered from protoplasts

The occurrence of genomic modifications in transgenic rice plants recovered from protoplasts and their transmission to the self-pollination progeny has been verfied with the random amplified polymorphic DNA (RAPD) approach. The plant was the Indica-type rice (Oryza sativa L.) cultivar Chinsurah Boro II. The analysed material was: (1) microspore-derived embryogenic rice cells grown in suspension culture, (2) transgenic plants recovered from protoplasts produced from the cultured cells and (3) the self-pollination progeny (two successive generations) of the transgenic plants. DNA purified from samples of these materials was PCR-amplified with different random oligonucleotide primers and the amplification products were analysed by agarose gel electrophoresis. Band polymorphism was scored and used in band-sharing analyses to produce a similarity matrix. Relationships among the analysed genomes were expressed in a dendrogram.The extensive DNA changes evidenced in cultured cells demonstrate the occurrence of somaclonal variation in the material used to produce protoplasts for gene transfer. Quantitatively reduced DNA changes were also found in the resulting transgenic plants and i their self-pollination progenies.While confirming the stability of the foreign gene in transgenic plants, this work gives molecular evidence for the occurrence of stable genomic changes in transgenic plants and points toin vitro cell culture as the causative agent. RAPDs are shown to be a convenient tool to detect and estimate the phenomenon at the molecular level. The methodology is also proposed as a fast tool to select those transgenic individuals that retain the most balanced genomic structure and to control the result of back-crosses planned to restore the original genome.     

水稻Ds插入纯合体的筛选和鉴定

采用Basta抗性鉴定、潮霉素抗性鉴定和PCR检测相结合的方法筛选和鉴定了水稻Ds插入纯合体。在T1代236个转化株系中,有16个株系的全部植株表现出对Basta的敏感,其余220个株系的植株表现出对Basta的抗性。经过3代的纯合筛选,共鉴定出Ds插入纯合体203个．这些Ds插入纯合体可用于构建Ac／Ds系统和对Ds插入突变体进行筛选和鉴定,为水稻功能基因组学研究提供了材料。     

Plant MITEs: useful tools for plant genetics and genomics

MTTEs (Miniature inverted-repeat transposabie elements) are reminiscence ot non-autonomous DNA (class Ⅱ) elements, which are distinguished from other transposable elements by their small size, short terminal inverted repeats (TIRs), high copy numbers, genie preference, and DNA sequence identity among family members. Although MITEs were first discovered in plants and still actively reshaping genomes, they have been isolated from a wide range of eukaryotic organisms. MITEs can be divided into Tourist-like, Stowaway-like, and pogo-like groups, according to similarities of their TIRs and TSDs (target site duplications). In despite of several models to explain the origin and amplification of MITEs, their mechanisms of transposition and accumulation in eukaryotic genomes remain poorly understood owing to insufficient experimental data. The unique properties of MITEs have been exploited as useful genetic tools for plant genome analysis. Utilization of MITEs as effective and informative genomic markers and pot     

Transgenic plants: performance,release and containment

This review focuses on transgenic plants, from the initial stages of the genetic modification process in the laboratory to their release stage in the field and indicates possible areas of concern and strategies for dealing with them. The classes of marker genes and issues about their safety, the gene flow and strategies that are used to isolate transgenic plants genetically are specifically examined. In addition, an assessment is provided of the phenomena which affect the performance of transgenic plants, such as gene disruption, the pleiotropic effect on plant phenotype and genetic variation. Finally, strategies are suggested for preventing unexpected consequences of transgenic plant production.The author is with the Department of Genetics, University of Leeds, Leeds LS2 9JT, UK     

Structural and functional analysis of rice genome

Rice is an excellent system for plant genomics as it represents a modest size genome of 430 Mb. It feeds more than half the population of the world. Draft sequences of the rice genome, derived by whole-genome shotgun approach at relatively low coverage (4-6 X), were published and the International Rice Genome Sequencing Project (IRGSP) declared high quality (> 10 X), genetically anchored, phase 2 level sequence in 2002. In addition, phase 3 level finished sequence of chromosomes 1, 4 and 10 (out of 12 chromosomes of rice) has already been reported by scientists from IRGSP consortium. Various estimates of genes in rice place the number at >50,000. Already, over 28,000 full-length cDNAs have been sequenced, most of which map to genetically anchored genome sequence. Such information is very useful in revealing novel features of macroand micro-level synteny of rice genome with other cereals. Microarray analysis is unraveling the identity of rice genes expressing in temporal and spatial manner and should help target candidate genes useful for improving traits of agronomic importance. Simultaneously, functional analysis of rice genome has been initiated by marker-based characterization of useful genes and employing functional knock-outs created by mutation or gene tagging. Integration of this enormous information is expected to catalyze tremendous activity on basic and applied aspects of rice genomics.