Characterization and diversity of novel PIF/Harbinger DNA transposons in Brassica genomes

331. Nouroz F, Noreen S, Heslop-Harrison J.S. 2016. Characterization and diversity of novel PIF/Harbinger DNA transposons in Brassica genomes. Pakistani Journal of Botany 48(1): 167-178. www.pakbs.org/pjbot/PDFs/48(1)/21.pdf

Among DNA transposons, PIF/Harbinger is most recently identified superfamily characterized by 3 bp target site duplications (TSDs), flanked by 14-45 bp terminal inverted repeats (TIRs) and displaying DDD or DDE domain displaying transposase. Their autonomous elements contain two open reading frames, ORF1 and ORF2 encoding superfamily specific transposase and DNA-binding domain. Harbinger DNA transposons are recently identified in few plants. In present study, computational and molecular approaches were used for the identification of 8 Harbinger transposons, of which only 2 were complete with putative transposase, while rest 6 lack transposase and are considered as defective or non-autonomous elements. They ranged in size from 0.5-4 kb with 3 bp TSDs, 15-42 bp TIRs and internal AT rich regions. The PCR amplification of Brassica Harbinger transposase revealed diversity and ancient nature of these elements. The amplification polymorphism of some non-autonomous Harbingers showed species specific distribution. Phylogenetic analyses of transposase clustered them into two clades (monocot and dicot) and five sub-clades. The Brassica, Arabidopsis and Malus transposase clustered into genera specific sub-clades; although a lot of homology in transposase was observed. The multiple sequence alignment of Brassica and related transposase showed homology in five conserved blocks. The DD35E triad and sequences showed similarity to already known Pong-like or Arabidopsis ATISI12 Harbinger transposase in contrast to other transposase having DD47E or DD48E motifs. The present study will be helpful in the characterization of Harbingers, their structural diversity in related genera and Harbinger based molecular markers for varietal/lines identification.

www.pakbs.org/pjbot/PDFs/48(1)/21.pdf

Locally archived at https://lra.le.ac.uk/handle/2381/37674

 

 

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Saffron crocus, cooking and Iran on the radio

crocus_saffron_pgiprotection.jpgThe spice Saffron is valued worldwide for its flavor, aroma and color. I’ve just broadcast an interview with Robin Young produced by Jill Ryan for NPR’s Here and Now program which let me tell you something about my enjoyment of saffron both as a consumer and scientist.

dsc01717paellaprawnssaffronnimesmarketSaffron is unusual in that it is equally at home in savory or sweet dishes, and as a drink made with hot water or cold vodka. Saffron is the most valuable agricultural product in the world: in a typical store, the weight of one penny (one cent piece, about 2.5g or a bit under a tenth of an ounce) will cost almost $25. That weight will come from about 200 flowers! But you only need a few strands per serving, costing a few cents – and most people who have enjoyed the real thing will be sure that it is worth it._phh5404saffron_rice.jpg

Saffron spice is the stigmas of the saffron Crocus flower: the three bright orange strands that normally capture the pollen. All _phh6001SaffronCrocusFlowerflowers have these stigmas, but the saffron crocus is unique in having stigmas with the sensory values. The whole flowers are picked, and the individual strands separated and dried. Different countries use slightly different drying temperatures and conditions, as well as soils and altitude, so saffron can vary in colour and aroma. Much is produced on family farms, and the whole family will be involved in picking from the early morning to drying later in the day. Most of the world’s saffron – something like 150 tons per year – is produced in Iran, where the climate and soils are favorable, as well as having people to pick by hand and separate the spice. Other saffron is produced in Kashmir (India/China/Pakistan border region), Greece and Spain, with smaller amounts from growers in many other countries.

Unfortunately, with such a high value product, fraud is an problem. Many types of fraud are possible: you can have the saffron repackaged so it looks like it comes from one country when it was actually produced elsewhere. Much worse though, is where the fraudsters copy the look or color. They will use things like corn silks or jute rope fibres, and colour from plants like safflower (a type of thistle) or tumeric (a type of ginger), or even synthetic dyes like Sudan yellow. As a consumer, you can look carefully at the saffron strands and check they look like the real thing. Smelling might be a problem where it is packaged, and shops would be less keen on tasting too, but you will soon recognize the flower petal-vanilla smell and tartly, slightly peppery taste. Be suspicious of buying powdered saffron, or if the price is too low: it can be kept for at least a couple of years, and nobody will pick hundreds of flowers for a few cents. Look for origin marks on packaging, and the reputation of the company selling it. We recently looked at 10 samples of saffron from supermarkets and those sold in small prepacks with clear labelling were all saffron. Those in packets with unclear labels or mentioning safflower and ‘saffron rice color’ had no aroma and were not saffron.dscn0071SaffronStigmasSpiceOnPaperLR

My scientific laboratory is part of the Saffronomics project. We are a consortium ranging from growers and producers through traders to scientists. As a group, we are developing ways to detect fraud in saffron, ranging from DNA and gene expression studies through to optical spectroscopy and isotope-ratio mass spectrometry. My own lab is interested in the diversity of the saffron crop – there is very little at the genetic level – and the origin from it’s closest wild relatives, some of the Crocuses which are often grown in gardens. The saffron crocus itself is sterile because it has three sets of chromosomes rather than the two of fertile plants and animals. We are also interested in development of high-value agricultural products and crops contributing to sustainable rural livelihoods. We have had several collaborations with researchers from Iran and I have much appreciated their input into our biodiversity and molecular genome studies. I am happy their farmers will have the chance now to export saffron to the US and hope development of this market will let new people enjoy the flavor and aroma of saffron.

LogoSaffronomics

 

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Preparation and fluorescent analysis of plant metaphase chromosomes

Chromosome Preparations: Schwarzacher 2016. www.molcyt.com

Chromosome Preparations: Schwarzacher 2016. http://www.molcyt.com

TS. Schwarzacher T. 2016. Preparation and fluorescent analysis of plant metaphase chromosomes. Chapter 7. In: Plant Cell Division: Methods and Protocols, ed Caillaud M-C; Methods in Molecular Biology 1370: 87-103. doi: 10.1007/978-1-4939-3142-2_7. Humana Press, Springer, New York. Local Copy: Chromosome Preparation. Trude Schwarzacher. www.molcyt.com

Good preparations are essential for informative analysis of both somatic and meiotic chromosomes, cytogenetics, and cell divisions. Fluorescent chromosome staining allows even small chromosomes to be visualized and counted, showing their morphology. Aneuploidies and polyploidies can be established for species, populations, or individuals while changes occurring in breeding lines during hybridization or tissue culture and transformation protocols can be assessed. The process of division can be followed during mitosis and meiosis including pairing and chiasma distribution, as well as DNA organization and structure during the evolution of chromosomes can be studied. This chapter presents protocols for pretreatment and fixation of material, including tips of how to grow plants to get good and healthy meristem with many divisions. The chromosome preparation technique is described using proteolytic enzymes, but acids can be used instead. Chromosome slide preparations are suitable for fluorochrome staining for fast screening (described in the chapter) or fluorescent in situ hybridization (see Schwarzacher and Heslop-Harrison, In situ hybridization. BIOS Scientific Publishers, Oxford, 2000).

KEYWORDS: Chromosome; DAPI; Fluorochromes; Heterochromatin; Meiosis; Metaphase; Proteolytic enzyme

Local Copy: Chromosome Preparation. Trude Schwarzacher. www.molcyt.com

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Saffron Crocus, quality and fraud in New York Times

_PHH2244SaffronStigmasPDOjarsProducts.jpgElaine Sciolino discusses saffron in the New York Times. Saffronomics partners Jean Thiercelin and Pat Heslop-Harrison are quoted, with the outcome of the project in developing methods to detect fraud and measure quality. In the article, the special qualities of saffron are discussed and many examples of the use in sweet and savoury foods explain the value in cooking. The high price encourages fraud and mis-labelling, but even at 20 Euro’s ($20, £15) a gram, the cost per serving is still not so great. Our work on the lack of genetic diversity in saffron and its relationships to wild crocus  was published in 2015 in Annals of Botany: this lack of diversity means that DNA-marker tests can be used to identify plant-based contamination but not the origin of saffron nor adulteration with dyes.

The full article is here http://mobile.nytimes.com/2015/12/30/dining/saffron-iran.html

 

 

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The molecular cytogenetic characterization of pistachio (Pistacia vera) suggests the arrest of recombination in the largest heteropycnotic pair HC1

Pistachio chromosomes: Sola-Campoy et al. 2015 PLoS One.

Pistachio chromosomes: Sola-Campoy et al. 2015 PLoS One.

TS. Sola-Campoy PJ, Robles F, Schwarzacher T, Ruiz Rejón C, de la Herrán R, Navajas-Pérez R (2015) The molecular cytogenetic characterization of Pistachio (Pistacia vera L.) suggests the arrest of recombination in the largest heteropycnotic pair HC1. PLoS ONE 10(12): e0143861. doi:10.1371/journal.pone.0143861. Local copy here.

This paper represents the first molecular cytogenetic characterization of the strictly dioecious pistachio tree (Pistacia vera L.). The karyotype was characterized by fluorescent in situ hybridization (FISH) with probes for 5S and 45S rDNAs, and the pistachio specific satellite DNAs PIVE-40, and PIVE-180, together with DAPI-staining. PIVE-180 has a monomeric unit of 176-178 bp and high sequence homology between family members; PIVE-40 has a 43 bp consensus monomeric unit, and is most likely arranged in higher order repeats (HORs) of two units. The P. vera genome is highly heterochromatic, and prominent DAPI positive blocks are detected in most chromosomes. Despite the difficulty in classifying chromosomes according to morphology, 10 out of 15 pairs (2n = 30) could be distinguished by their unique banding patterns using a combination of FISH probes. Significantly, the largest pair, designated HC1, is strongly heteropycnotic, shows differential condensation, and has massive enrichment in PIVE-40 repeats. There are two types of HC1 chromosomes (type-I and type-II) with differing PIVE-40 hybridization signal. Only type-I/II heterozygotes and type-I homozygotes individuals were found. We speculate that the differentiation between the two HC1 chromosomes is due to suppression of homologous recombination at meiosis, reinforced by the presence of PIVE-40 HORs and differences in PIVE-40 abundance. This would be compatible with a ZW sex-determination system in the pistachio tree.

 

TS. Sola-Campoy PJ, Robles F, Schwarzacher T, Ruiz Rejón C, de la Herrán R, Navajas-Pérez R (2015) The molecular cytogenetic characterization of Pistachio (Pistacia vera L.) suggests the arrest of recombination in the largest heteropycnotic pair HC1. PLoS ONE 10(12): e0143861. doi:10.1371/journal.pone.0143861. Local copy here.

 

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Banana research at the botanic garden

Banana fruit bunch with male flower

Banana fruit bunch with male flower

324. Heslop-Harrison, P. 2015. Banana research at the botanic garden. University of Leicester Botanic Garden Newsletter 12: 4. November 2015.

Dessert bananas and the cooking bananas or plantains are among the oldest crops in the world. Most crops were domesticated through a long pathway of selection and crossing but, for banana, virtually all the two thousand varieties which are grown throughout the tropics were collected as spontaneous mutants in the wild with the extraordinary property of having large fleshy fruit without any seeds. Most varieties of banana are also unusual in having three sets of chromosomes, a condition known as triploid.

The wild progenitors of the domesticated banana are from south-east Asia and are currently known as Musa acuminata (with a genome designated as ‘A’, green on the map below) and Musa balbisiana (with a ‘B’ genome, orange on the map below). About 15% of the world’s banana production is for the export trade, and is based on a single variety, ‘Cavendish’. This sweet banana has the genome constitution AAA. Banana varieties that are hybrids with AAB and ABB genome constitutions are a staple food for a billion people in Asia and Africa, and in Leicester we are fortunate that many of these plantains and cooking bananas (eaten fried or steamed and mashed as a vegetable) are easily available in the market and speciality shops.

In many parts of Asia, banana leaves are also used as disposable plates or as wrappers for steaming rice or banana fruit. Interestingly, there is now the possibility that these two currently recognized A and B species will be merged into one; this would probably mean resurrecting one of the original names given by Linneaus, Musa paradisiaca L. or Musa sapientum L.

Traditionally, bananas are propagated by side suckers to the main stem but, in commercial plantations, most plants are now multiplied through tissue culture to ensure disease-free planting material. In our research group, we study the diversity and evolution of bananas at the DNA level, and are looking for new diversity to improve current varieties. The amount of DNA in plant genomes varies widely, from less than 100 Mbp in some carnivorous plants such as Genlisea, to more than 17,000 Mbp in wheat and pines. Bananas are at the lower end of the range, with about 550 Mbp. We were involved in the international consortium that sequenced all the DNA in banana in 2012, and thus we have a reference for all the genes and regulatory sequences present in the species.

This is vital if breeding programmes are to produce new disease-resistant varieties. Disease is a major problem in banana production in the topics: bananas suffer from fungi, bacteria, viruses, insects and nematodes. About a third of the cost of growing the bananas is spent on crop protection chemicals, and all fruit bunches are covered with blue bags to deter insects from damaging the fruit and allowing entry of secondary fungal. Eradication of disease in some areas costs millions of pounds and requires destruction of millions of plants. Very careful agronomy can control diseases, such as burning plants at the first sign of infection, or dipping machetes in bleach between harvesting each plant, but such measures required huge amounts of training and labour. Some diseases cannot be controlled with agronomy or chemicals, and means banana production is lost to an area. Our current work is looking at the diversity present in the whole banana genome at various positions related to diseases, we are also looking how the variation can be exploited in generating new varieties of banana which are disease resistant.

324. Heslop-Harrison, P. 2015. Banana research at the botanic garden. University of Leicester Botanic Garden Newsletter 12: 4. November 2015.

 

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Repetitive DNA in eukaryotic genomes

Biscotti et al. 2015. Repetitive DNA in the eukaryotic genome. Chromosome Research

Biscotti et al. 2015. Repetitive DNA in the eukaryotic genome. Chromosome Research

322. Biscotti MA, Olmo E, Heslop-Harrison JS. 2015. Repetitive DNA in eukaryotic genomes. Chromosome Research 23(3): 415-420. DOI: 10.1007/s10577-015-9499-z

Biscotti Repetitive DNA Author Version

Link to Repetitive DNA summary Diagram Molcyt.com DNA

Repetitive DNA — sequence motifs repeated hundreds or thousands of times in the genome— makes up the major proportion of all the nuclear DNA in most eukaryotic genomes. However, the significance of repetitive DNA in the genome is not completely understood, and it has been considered to have both structural and functional roles, or perhaps even no essential role. High-throughput DNA sequencing reveals huge numbers of repetitive sequences. Most bioinformatic studies focus on low-copy DNA including genes, and hence, the analyses collapse repeats in assemblies presenting only one or a few copies, often masking out and ignoring them in both DNA and RNA read data. Chromosomal studies are proving vital to examine the distribution and evolution of sequences because of the challenges of analysis of sequence data. Many questions are open about the origin, evolutionary mode and functions that repetitive sequences might have in the genome. Some, the satellite DNAs, are present in long arrays of similar motifs at a small number of sites, while others, particularly the transposable elements (DNA transposons and retrotranposons), are dispersed over regions of the genome; in both cases, sequence motifs may be located at relatively specific chromosome domains such as centromeres or subtelomeric regions. Here, we overview a range of works involving detailed characterization of the nature of all types of repetitive sequences, in particular their organization, abundance, chromosome localization, variation in sequence within and between chromosomes, and, importantly, the investigation of their transcription or expression activity. Comparison of the nature and locations of sequences between more, and less, related species is providing extensive information about their evolution and amplification. Some repetitive sequences are extremely well conserved between species, while others are among the most variable, defining differences between even closely relative species. These data suggest contrasting modes of evolution of repetitive DNA of different types, including selfish sequences that propagate themselves and may even be transferred horizontally between species rather than by descent, through to sequences that have a tendency to amplification because of their sequence motifs, to those that have structural significance because of their bulk rather than precise sequence. Functional consequences of repeats include generation of variability by movement and insertion in the genome (giving useful genetic markers), the definition of centromeres, expression under stress conditions and regulation of gene expression via RNA moieties. Molecular cytogenetics and bioinformatic studies in a comparative context are now enabling understanding of the nature and behaviour of this major genomic component.

repetitive DNA, tandem repeats, genomics, junk DNA, transposons, satellite DNA, retrotransposons, review

Biscotti MA, Olmo E, Heslop-Harrison JS. 2015. Repetitive DNA in eukaryotic genomes. Chromosome Research 23(3): 415-420. DOI: 10.1007/s10577-015-9499-z

Biscotti Repetitive DNA Author Version

Link to Repetitive DNA summary Diagram Molcyt.com DNA

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