Miniature inverted-repeat transposable elements (MITEs) are a special type of Class 2 non-autonomous transposable element (TE) that are abundant in the non-coding regions of the genes of many plant and animal species. The accurate identification of MITEs has been a challenge for existing programs because they lack coding sequences and, as such, evolve very rapidly. Because of their importance to gene and genome evolution, we developed MITE-Hunter, a program pipeline that can identify MITEs as well as other small Class 2 non-autonomous TEs from genomic DNA data sets. The output of MITE-Hunter is composed of consensus TE sequences grouped into families that can be used as a library file for homology-based TE detection programs such as RepeatMasker. MITE-Hunter was evaluated by searching the rice genomic database and comparing the output with known rice TEs. It discovered most of the previously reported rice MITEs (97.6%), and found sixteen new elements. MITE-Hunter was also compared with two other MITE discovery programs, FINDMITE and MUST. Unlike MITE-Hunter, neither of these programs can search large genomic data sets including whole genome sequences. More importantly, MITE-Hunter is significantly more accurate than either FINDMITE or MUST as the vast majority of their outputs are false-positives.

/pdf/mite-hunter-a-program-for-discovering-miniature-inverted-athndhakjl.pdf

MITE-Hunter: a program for discovering miniature inverted-repeat transposable elements from genomic sequences

A Joint Announcement on Genome Sequence Standards Genome project standards in a new era of sequencing P. S. G. Chain 1,2,3,4,22,* , D. V. Grafham 5,* , R. S. Fulton 6 , M. G. FitzGerald 7 , J. Hostetler 8 , D. Muzny 9 , J. C. Detter 1,10 , J. Ali 11 , B.Birren 7 , D. C. Bruce 1, 10 , C. Buhay 9 , J. R. Cole 3,4 , Y. Ding 9 , S. Dugan 9 , D. Field 12 , G. M. Garrity 3,4 , R. Gibbs 9 , T. Graves 6 , C. S. Han 1, 10 , S. H. Harrison 3 , S. Highlander 9 , P. Hugenholtz 1 , H. M. Khouri 13 , C. D. Kodira 7,23 , E. Kolker 14,15 , N. C. Kyrpides 1 , D. Lang 1,2 , A. Lapidus 1 , S. A. Malfatti 1,2 , V. Markowitz 16 , T. Metha 7 , K. E. Nelson 8 , J. Parkhill 5 , S. Pitluck 1 , X. Qin 9 , T. D. Read 17 , J. Schmutz 18 , S. Sozhamannan 19 , R. Strausberg 8 , G. Sutton 8 , N. R. Thomson 5 , J. M. Tiedje 3,4 , G. Weinstock 6 , A. Wollam 6 , and the entire GSC 20 and HMP Jumpstart 21 consortia. U.S. Department of Energy Joint Genome Institute, Walnut Creek, California 94598, USA Biosciences and Biotechnology Division, Lawrence Livermore National Laboratory, Livermore, California 94550, USA Microbiology & Molecular Genetics, Michigan State University, East Lansing, Michigan 48824, USA Center for Microbial Ecology, Michigan State University, East Lansing, Michigan 48824, USA The Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge CB10 1SA, United Kingdom The Genome Center, Washington University School of Medicine, St Louis, Missouri 63108, USA The Broad Institute of Massachusetts Institute of Technology and Harvard, Cambridge, Massachusetts 02141, USA J. Craig Venter Institute, Rockville, Maryland 20850, USA Human Genome Sequencing Center, Baylor College of Medicine, Houston, Texas 77030, USA Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USA Ontario Institute for Cancer Research, Toronto, Ontario M5G 0A3, Canada Natural Environmental Research Council Centre for Ecology and Hydrology, Oxford, Oxfordshire OX1 3SR, UK National Center for Biotechnology Information, National Library of Medicine, Rockville, Maryland 20850, USA Seattle Children’s Hospital and Research Institute, Seattle, Washington 98101, USA Biomedical & Health Informatics Division, MEBI, University of Washington School of Medicine, Seattle, Washington 98195, USA Biological Data Management and Technology Center, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA Emory GRA Genomics Core, Emory University School of Medicine, Atlanta, Georgia 30322, USA HudsonAlpha Genome Sequencing Center, HudsonAlpha Institute, Huntsville, Alabama 35806, USA Biological Defense Research Directorate, Naval Medical Research Center, Silver Spring, Maryland 20910, USA Genomic Standards Consortium Human Microbiome Project Jumpstart Consortium Current address: Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USA Current address: 454 Life Sciences, Branford, Connecticut 06405, USA *Address correspondence to Patrick Chain (pchain@lanl.gov) and Darren Grafham (dg1@sanger.ac.uk)

Genome Project Standards in a New Era of Sequencing

Class 2 transposable elements (TEs) are the predominant elements in and around plant genes where they generate significant allelic diversity. Using the complete sequences of four grasses, we have performed a novel comparative analysis of class 2 TEs. To ensure consistent comparative analyses, we re-annotated class 2 TEs in Brachypodium distachyon, Oryza sativa (rice), Sorghum bicolor and Zea mays and assigned them to one of the five cut-and-paste superfamilies found in plant genomes (Tc1/mariner, PIF/Harbinger, hAT, Mutator, CACTA). We have focused on noncoding elements because of their abundance, and compared superfamily copy number, size and genomic distribution as well as correlation with the level of nearby gene expression. Our comparison revealed both unique and conserved features. First, the average length or size distribution of elements in each superfamily is largely conserved, with the shortest always being Tc1/mariner elements, followed by PIF/Harbinger, hAT, Mutator and CACTA. This order also holds for the ratio of the copy numbers of noncoding to coding elements. Second, with the exception of CACTAs, noncoding TEs are enriched within and flanking genes, where they display conserved distribution patterns, having the highest peak in the promoter region. Finally, our analysis of microarray data revealed that genes associated with Tc1/mariner and PIF/Harbinger noncoding elements have significantly higher expression levels than genes without class 2 TEs. In contrast, genes with CACTA elements have significantly lower expression than genes without class 2 TEs. We have achieved the most comprehensive annotation of class 2 TEs to date in these four grass genomes. Comparative analysis of this robust dataset led to the identification of several previously unknown features of each superfamily related to copy number, element size, genomic distribution and correlation with the expression levels of nearby genes. These results highlight the importance of distinguishing TE superfamilies when assessing their impact on gene and genome evolution.

/pdf/comparison-of-class-2-transposable-elements-at-superfamily-4llu6l7zmt.pdf

Comparison of class 2 transposable elements at superfamily resolution reveals conserved and distinct features in cereal grass genomes.

Miniature inverted-repeat transposable elements (MITEs) are a particular type of defective class II transposons present in genomes as highly homogeneous populations of small elements. Their high copy number and close association to genes make their potential impact on gene evolution particularly relevant. Here, we present a detailed analysis of the MITE families directly related to grapevine “cut-and-paste” transposons. Our results show that grapevine MITEs have transduplicated and amplified genomic sequences, including gene sequences and fragments of other mobile elements. Our results also show that although some of the MITE families were already present in the ancestor of the European and American Vitis wild species, they have been amplified and have been actively transposing accompanying grapevine domestication and breeding. We show that MITEs are abundant in grapevine and some of them are frequently inserted within the untranslated regions of grapevine genes. MITE insertions are highly polymorphic among grapevine cultivars, which frequently generate transcript variability. The data presented here show that MITEs have greatly contributed to the grapevine genetic diversity which has been used for grapevine domestication and breeding.

/pdf/recent-amplification-and-impact-of-mites-on-the-genome-of-48dgd2gaah.pdf

Recent amplification and impact of MITEs on the genome of grapevine (Vitis vinifera L.)

We characterized a Brassica miniature inverted repeat transposable element (MITE) from the Stowaway superfamily, designated BraSto (Bra
 ssica
 Sto
 waway). BraSto copy number was assessed using real-time quantitative PCR in the two diploid species B. rapa (genome A) and B. oleracea (genome C) and the corresponding allotetraploid species B. napus (genome AC). Phylogenetic relationships among a set of 131 BraSto copies were then analyzed. BraSto appears to have been only moderately amplified in the Brassica genome and was still active recently with marks of proliferation in both diploid Brassica species, which diverged 3.75 million years ago, but also in the allotetraploid species after reuniting of the two differentiated genomes. We characterized insertion sites for low-divergence BraSto copies among the gene space of the B. rapa genome using bioinformatics approaches. For BraSto copies localized nearby or within genes, we observed frequent associations of BraSto with putative promoters and regulatory regions of genes, but exclusion from coding regions. In addition, BraSto was significantly similar to several Brassica expressed sequence tags (ESTs), including stress-induced ESTs. We also demonstrated the enrichment of BraSto sequences in binding sites for transcription factors and other regulatory elements. Our results lead to the question of a role for BraSto in the regulation of gene expression: this putative role, if further confirmed experimentally, would help to obtain a new insight into the significance of MITEs in the functional plant genome.

BraSto, a Stowaway MITE from Brassica: recently active copies preferentially accumulate in the gene space.

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.

hATpin, a family of MITE-like hAT mobile elements conserved in diverse plant species that forms highly stable secondary structures.

Bacterial contamination represents a serious problem for plant tissue culture research and applications. Bacterial interference with normal plant physiology and morphology can generate misleading conclusions if the presence of bacteria is ignored. Bacterial contaminants in in vitro plant culture are typically detected by direct observation; thus, it is assumed that cultures without visible symptoms are bacteria free. Here, we demonstrate that contaminating Bacillus DNA in plant DNA solutions from asymptomatic plants can interfere with the analysis of somaclonal variation in chrysanthemum. We studied somaclonal variation in chrysanthemum using short semi-specific PCR primers based on conserved motifs in NBS–LRR disease resistance genes and in mobile elements. Instead of true somaclonal variation we found three polymorphic bands derived from contaminant bacterial DNA in plant extracts. Although the detection of asymptomatic bacteria in in vitro plant cultures is a major issue, we found that it has not been adequately addressed to date, particularly for studies on somaclonal variation. We reviewed the most commonly cited contaminant bacteria in in vitro plant culture and designed specific 16S rRNA gene-based PCR primers for the main genera causing contamination (Bacillus, Pseudomonas, Staphylococcus, Lactobacillus, Erwinia/Enterobacter
 and
 Xanthomonas). Using a panel of pure bacterial DNAs, artificial mixes of bacterial/plant DNAs, and in vitro plant cultures with and without visible contamination we demonstrated that our primers are in most instances both reliable and sensitive, and appropriate for the identification and tracking of the most frequent bacterial contaminants in plant in vitro cultures. Implications of bacterial identification to molecular analysis of somaclonal variation and plant culture decontamination are discussed.

Bacterial contamination of in vitro plant cultures: confounding effects on somaclonal variation and detection of contamination in plant tissues

Improving the performance of peach varieties in the context of climate change requires multiple approaches. Not only will climate change alter plant phenology, but it will also drive negative effects of several biotic and abiotic stressors. The challenge is to improve adaptation of varieties to a changing environment, while maintaining organoleptic qualities of the fruit. This chapter focuses on the progress in genomics-assisted breeding in peach to break barriers in conventional breeding. Breeding climate-smart (CS) peach trees requires the identification of CS traits used in the adaptation to high levels of temperature, CO2, water deprivation and biotic stress. Relevant CS traits, such as those that control flowering time (chilling and heat requirements), biotic and abiotic stress tolerance (pests and diseases; water-nutrient efficiency), require prioritization. Here, we review classical mapping and breeding of peach varieties, the progress and limitations of the used of marker-assisted selection and breeding (MAS and MAB, respectively) in expression of traits, such as fruit quality and stress tolerance, and describe the rationale for the use of molecular breeding. Genetic diversity analysis of Prunus germplasm, genome-wide association mapping and molecular mapping of CS genes and QTLs, and MAB and genomics-aided breeding for CS traits are also reviewed. MAS and MAB have previously been considered as the optimal solution to plant breeding in the genomics era of plant biology, but genomic selection currently presents a promising alternative. Genomic selection is a marker-based strategy that accounts for quantitative traits controlled by a large number of genes with small effects like many CS traits. The precise phenotypic assessment and appropriate biometric analysis used to identify genotype responses is also discussed. The small, but active international peach research community has delivered a high-quality sequenced and annotated genome, along with several genomic tools, that potentiate each other in a positive feedback. Bioinformatics and computational biology are at the current a forefront of plant breeding programs and deal with diverse functional genomics datasets of gene expression, metabolomics and physiological stress responses. Taken together, existing genomic knowledge and tools may be used to confront the challenges of the development of peach varieties adapted to changing climate scenarios.

Santiago Moreno-Vázquez

Papers

hATpin, a family of MITE-like hAT mobile elements conserved in diverse plant species that forms highly stable secondary structures.

Bacterial contamination of in vitro plant cultures: confounding effects on somaclonal variation and detection of contamination in plant tissues

Genomic-Based Breeding for Climate-Smart Peach Varieties

Quantitative evaluation of Heterodera avenae females in soil and root extracts by digital image analysis