Wordle of the blog

The Wordle (http://www.wordle.net/) of the blog. This utility generates word clouds from the text that is inserted. In this case the whole blog. Unfortunately I've had little time to update the blog recently. I will try to catch up soon. Sorry....

Software evaluation - FlowJo

Following this year's ISAC congress I've been aware of many commercial software packages for the analysis of flow cytometric data. After attending to the FlowJo's commercial tutorial I was very curious to test it in the analysis of plant DNA flow cytometry results. But, what advantages can this powerful package (it was designed to ease the analysis of multiparametric results, besides other things) have in the analysis of simple, one to two parameters results, as it is the analysis of DNA histograms obtained from the isolation of plant nuclei? That is to say, which positive features FlowJo has that are not present in the software packages that are provided with your flow cytometer and that may improve your daily analysis of the obtained results?

Having used only a few of its huge list of features, I highlight two of them: batch operations (repetitive analyses performed on multiple samples) and output generation (e.g. easy exportation of tabular data to an excel file, for example). Both these features enable a much faster and easier analysis of the obtained results. For example, you can create the necessary regions and gating in a specific histogram (file) and then just copy those settings to all the other ones. I.e. to say that it is very easy to apply common procedures to all the files of an experiment; you should only do slight adjustments, but even this can be done in a very quick and efficient way. After all the analysis is done it is also very easy to export your results in the form of a table (among other options) to a variety of external platforms, let's say Excel, so that you can make further analysis. You can also create a workspace that contains all the files of your experiment and within the workspace several groups that may have specific assays. This enables the fast and easy organization of your files. Another interesting feature for the plant DNA flow cytometry community is the cell cycle analysis procedure. However, I've still not been able to test it appropriately.

The biggest disadvantage that I can point out is the price of FlowJo. Despite that it can be quite acceptable for the users from the clinical area, I find that it is still quite high for the users in plant flow cytometry. Another minor negative point is the somehow stylized presentation of the DNA histograms, that sometimes seem slightly better than they were when obtained using the standard software of the flow cytometer. However, the enormous array of available statistics provide you the real values (e.g. CV values) of your data.

So far, my experience has been very good and I personally recommend you to download a fully functional 30 day trial version of the software. It's quite straightforward to use, despite that I must confess that the tutorial that I attended in the ISAC's Congress proved to be very useful for a faster start. I would say that there's nothing better than to try it yourself.

The European Cytometry Network webpage

The webpage of the recently created European Cytometry Network (for further info click here) has been in beta testing already for several months. This may be a very useful and attractive platform created as a social network for the easy exchange of ideas and information between European scientists interested on cytometry. There's a special group exclusively dedicated to plant flow cytometry, which has already 15 members from several areas of plant sciences. So, it may be another web platform (together with the forum) for the discussion of general topics in plant flow cytometry as well as the place to pose any question or doubt related with the cytometric applications in plant sciences.

As the network works as invitation only, in case you are interested in joining us, please just send me an e-mail that I will promptly reply to you.

Make a good profit of it... it was created for you.

List of recently published papers on plant flow cytometry - June, July and August

Exceptionally, due to my recent busy times exposed in a previous post, this highlight of the recently published articles on Plant Flow Cytometry will include the months of June, July and August.

Genome size:
Thompson JN, Merg KF. Evolution of polyploidy and the diversification of plant-pollinator interactions. Ecology (2008) 89:2197-2206.

de Abreu IS, Carvalho CR, Clarindo WR. Chromosomal DNA content of sweet pepper determined by association of cytogenetic and cytometric tools. Plant Cell Reports (2008) 27:1227-1233.

Brito G, Loureiro J, Lopes T, Rodriguez E, Santos C. Genetic characterisation of olive trees from Madeira Archipelago using flow cytometry and microsatellite markers. Genetic Resources and Crop Evolution (2008) 55:657-664.

Consaul LL, Gillespie LJ, Waterway MJ. Systematics of North American Arctic diploid Puccinellia (Poaceae): Morphology, DNA content, and AFLP markers. Systematic Botany (2008) 33:251-261.

Garcia S, Canela MA, Garnatje T, Mcarthur ED, Pellicer J, Sanderson SC, Valles J. Evolutionary and ecological implications of genome size in the North American endemic sagebrushes and allies (Artemisia, Asteraceae). Biological Journal of the Linnean Society (2008) 94:631-649.

Stahlberg D, Hedren M. Systematics and phylogeography of the Dactylorhiza maculata complex (Orchidaceae) in Scandinavia: insights from cytological, morphological and molecular data. Plant Systematics and Evolution (2008) 273:107-132.

Eilam T, Anikster Y, Millet E, Manisterski J, Feldman M. Nuclear DNA amount and genome downsizing in natural and synthetic allopolyploids of the genera Aegilops and Triticum. Genome (2008) 51:616–627.

Ploidy level:
Biotechnology
Izumikawa Y, Takei S, Nakamura I, Mii M. Production and characterization of inter-sectional hybrids between Kalanchoe spathulata and K. laxiflora (= Bryophyllum crenatum). Euphytica (2008) 163:123-130.

Skrlep K, Bergant M, De Winter GM, Bohanec B, Zel J, Verpoorte R, Van Iren F, Camloh M. Cryopreservation of cell suspension cultures of Taxus x media and Taxus floridana. Biologia Plantarum (2008) 52:329-333.

Jin SX, Mushke R, Zhu HG, Tu LL, Lin ZX, Zhang YX, Zhang XL. Detection of somaclonal variation of cotton (Gossypium hirsutum) using cytogenetics, flow cytometry and molecular markers. Plant Cell Reports (2008) 27:1303-1316.

Yang XM, An LZ, Xiong YC, Zhang JP, Li Y, Xu SJ. Somatic embryogenesis from immature zygotic embryos and monitoring the genetic fidelity of regenerated plants in grapevine. Biologia Plantarum (2008) 52:209-214

Zdravkovic-Korac S, Calic-Dragosavac D, Uzelac B, Janosevic D, Budimir S, Vinterhalter B, Vinterhalter D. Secondary somatic embryogenesis versus caulogenesis from somatic embryos of Aesculus carnea Hayne.: developmental stage impact. Plant Cell Tissue and Organ Culture (2008) 94:225-231.

Biosystematics
Lubinsky P, Cameron KM, Molina MC, Wong M, Lepers-Andrzejewski S, Gomez-Pompa A, Kim SC. Neotropical roots of a Polynesian spice: The hybrid origin of Tahitian vanilla, Vanilla tahitensis (Orchidaceae). American Journal of Botany (2008) 95:1040-1047.

Mereda P, Hodalova I, Martonfi P, Kucera J, Lihova J. Intraspecific variation in Viola suavis in Europe: Parallel evolution of white-flowered morphotypes. Annals of Botany (2008) 102:443-462.

Schenk MF, Thienpont CN, Koopman WJM, Gilissen LJWJ, Smulders MJM. Phylogenetic relationships in Betula (Betulaceae) based on AFLP markers. Tree Genetics & Genomes (2008) 4:911-924.

Apomixis
Siena LA, Sartor ME, Espinoza F, Quarin CL, Ortiz JPA. Genetic and embryological evidences of apomixis at the diploid level in Paspalum rufum support recurrent auto-polyploidization in the species. Sexual Plant Reproduction (2008) 21:205-215.

Plant breeding
Martinez-Reyna JM, Vogel KP. Heterosis in switchgrass: Spaced plants. Crop Science (2008) 48:1312-1320.

Endoreduplication:
Teyssier E, Bernacchia G, Maury S, Kit AH, Stammitti-Bert L, Rolin D, Gallusci P. Tissue dependent variations of DNA methylation and endoreduplication levels during tomato fruit development and ripening. Planta (2008) 228:391-399.

Cell Cycle:
Da Silva EAA, Toorop PE, Van Lammeren AAM, Hilhorst HWM. ABA inhibits embryo cell expansion and early cell division events during coffee (Coffea arabica 'Rubi') seed germination. Annals of Botany (2008) 102:425-433.

Short resume of the Xth Symposium of the International Organization of Plant Biosystematics

Before updating all the usual sections of the blog, I want to leave you a very short overview of the Xth Symposium of the International Organization of Plant Biosystematics. Overall, the Symposium was a success both in organization and scientific value. The venue was held on the beautiful High Tatras which was another positive aspect.

Flow cytometry speaking it was nice to see that there are more and more researchers using this technique to help solving byosystematic and taxonomy problems. The number of posters where flow cytometry was used exceeded the 20 and several talks also referred its usefulness and potential for large scale ploidy level and genome size analyses. The only regret was the unexpected cancelling of D. Soltis talk due to passport issuing problems. Maybe next time...

Finally, Jan Suda wonderfully and enthusiastically presented a talk addressing all the potentialities that the technique has in the field of Biosystematics, which was a sure highlight of the congress's second day.

I leave you some other pictures from the Tatras journey:

Xth Symposium of the International Organization of Plant Biosystematics

The next Symposium of the International Organization of Plant Biosystematics will occur next week (from the 2nd to the 4th of July) in Visoké Tatry, Slovakia. This year the meeting will be devoted to the evolution of plants in montainous and alpine habitats. The impact that flow cytometry has been having in this area is well patent in the Symposium Scientific Programme as there will be one talk totally dedicated to it (Jan Suda, Praha - From individuals to populations: the impact of flow cytometry on understanding polyploid evolution in mountain plants) and some more talks that will certainly focus on data obtained using this powerful and high throughput technique.

The main topics that will be focused are:
PHYLOGEOGRAPHY
BIOGEOGRAPHY
EVOLUTIONARY PROCESSES IN EUROPEAN MOUNTAIN RANGES
POLYPLOIDY
MOLECULAR APPROACHES IN PLANT EVOLUTION
ECOLOGICAL FACTORS IN PLANT EVOLUTION
EVOLUTIONARY PROCESSES IN EXTRA-EUROPEAN MOUNTAINS
ROLE OF APOMIXIS IN PLANT EVOLUTION

There is still the possibility to register on site, so do not miss this opportunity to, besides enjoying a Symposium with a rather interesting programme (which should be the main purpose!), discover this beautiful mountain area of Slovakia.

I hope to see you there.

Plant Flow Cytometry - Far Beyond the Stone Age Commentary on Cytometry Journal

In a close association with the review of Sergio Ochatt on the application of flow cytometry in plant breeding (see previous post) that appears in the current issue of Cytometry Part A journal we have been invited to write a commentary on the current state of flow cytometry in plant sciences. It is a short overview of the applications of flow cytometry to analyse plant cells and once again the blog, forum and FLOWer database were not forgotten. We hope that you enjoy the reading.

You can download it for free in here: Plant Flow Cytometry - Far Beyond the Stone Age

The first reference in a manuscript to the Plant FCM blog and forum

In the last issue of Cytometry a review on the application of flow cytometry to plant breeding is presented by Sergio J. Ochatt from INRA (Dijon Cedex, France). The manuscript explores some of the interesting uses of flow cytometry in the area of plant breeding and, by the end, in an overview of the progress in recent years, both this blog and the plant flow cytometry forum are referred with the respective web address. We are very thankful for this mention and it is already a victory that we are slowly gaining some visibility within the plant FCM community.

Thank you all for being so supportive.

ISAC Congress - Plant FCM Workshop Presentations

Despite that the Congress was already over almost one month ago, we are still summarizing some of the plant related events. Now, it is time to provide you with the presentations given in the plant FCM workshops (click on each image for download). The presentations may be used without copyright restrictions except for commercial or for profit use, provided there is some credit to the author and reference to this blog.

Workshop - Plant genome structure and gene expression - organized by David Galbraith and Jaroslav Dolezel











Presentations available by Jan Suda (Preparation and storage of plant samples for DNA flow cytometry), João Loureiro (The effect of cytosol on quantitative staining of nuclear DNA), Johann Greilhuber (Standardization and Standards), Jaroslav Dolezel (Chromosome analysis and sorting) and David Galbraith (Analysis of gene expression, including flow analysis and sorting of organelles and large cells)

Workshop - Plant evolutionary biology, biosystematics and ecology - organized by Brian Husband and Jan Suda







Presentations available by Jan Suda (The impact of FCM on plant Biosystematics and Taxonomy), Paul Kron (Flow Cytometry: Applications in the Study of Plant Reproductive Systems) and Brian Husband (Flow Cytometry and The Evolutionary Dynamics of Polyploids).

We hope that you enjoy the presentations.

List of recently published papers on plant flow cytometry - May

Below is the list of papers that came to our hands/knowledge during the month of April. In here there are already included some of the contributions made through the FLOWer database webpage. Many thanks for the authors that contributed with their work.

Genome size:
Carvalho CR, Clarindo WE, Praça MM, Araújo FS, Carels N. Genome size, base composition and karyotype of Jatropha curcas L., an important biofuel plant. Plant Science (2008) 174:613-617.

Rossi AAB, Clarindo WR, Carvalho CR, Oliveira LO. Karyotype and nuclear DNA content of Psychotria ipecacuanha: a medicinal species. Cytologia 73 (2008):53–60.

Ecology
Ayres DR, Grotkopp E, Zaremba K, Sloop CM, Blum MJ, Bailey JP, Anttila CK, Strong DR. Hybridization between invasive Spartina densiflora (Poaceae) and native S. foliosa in San Francisco Bay, California, USA. American Journal of Botany (2008) 95:713-719.

Biotechnology
Makowczynska J, Andrzejewska-Golec E, Sliwinska E. Nuclear DNA content in different plant materials of Plantago asiatica L. cultured in vitro. Plant Cell, Tissue and Organ Culture (2008) 94:65–71

Ploidy level:
Biotechnology
Kantartzi S, Roupakias DGProduction of aneuploids of the cotton hybrid G. barbadense x G. hirsutum L. via intergeneric pollination with Abelmoschus esculentus. Euphytica (2008) 161:319-327.

Orbovic V, Calovic M, Viloria Z, Nielsen B, Gmitter FG, Castle WS, Grosser JW. Analysis of genetic variability in various tissue culture-derived lemon plant populations using RAPD and flow cytometry. Euphytica (2008) 161:329-335.

Guillemin ML, Faugeron S, Destombe C, Viard F, Correa JA, Valero M. Genetic variation in wild and cultivated populations of the haploid-diploid red alga Gracilaria chilensis: how farming practices favor asexual reproduction and heterozygosity. Evolution (2008) 62-6: 1500–1519

Functional cytometry:
Apoptosis
Tilbrook J, Tyerman SD. Cell death in grape berries: varietal differences linked to xylem pressure and berry weight loss. Functional Plant Biology (2008) 35:173-184.

FLOWer database version 1.0 has just been launched

I am very pleased to announce that version 1.0 of the FLOWer database is finally ready and online (http://flower.web.ua.pt/). After some serious problems with the programming of the previous version of the database, we are very glad that the first release of the database is up and running. You are welcome to visit it and I encourage you to use the database as a unique resource of publications on DNA flow cytometry in plant sciences. Your opinion is very important to us, so it would be nice to receive any feedback through the shoutbox available in the home page or directly to this e-mail. The database presently harbours 826 publications, and you are welcome to provide us with any missing ones through the contribution area available in the menu bar of the home page.

This is part of the e-mail that was sent to many colleagues of the plant flow cytometry area, where this blog and the forum of plant flow cytometry were also promoted. I hope that you like all these resources that were made available to you.

Many thanks for stopping by.

Some other ISAC posters related with plant flow cytometry

After having talked with some of the participants related with plant flow cytometry, some of them agreed in sending me a jpg version of their posters. Please click on each image for the full size of the poster. As soon as I receive more posters, I will update this post.

Contributors:
Tomas Urfus - Department of Botany, Charles University in Prague, Czech Republic.
Sara V. Petersson - Umea Plant Science Center, Department of Forest Genetics and Plant Physiology, Swedish University of Agricultural Sciences, Umea, Sweden.
Jingly Zhang - HortResearch, New Zealand.

Many thanks to all the contributors.

First review to the book Flow Cytometry with Plant Cells

The last issue of Cytometry part A (Volume 73A Issue 6) has just published the first review to the book Flow Cytometry with Plant Cells: Analysis of Genes, Chromosomes and Genomes that was launched last year (see the side bar for full details). The review was made by Dr. Sergio Ochatt from the INRA, Dijon, France and the author carefully summarized what the book has to offer.You can download it for free by clicking this link.


As an author of one chapter I am very happy to finally see a review to the book. I am positive that it will be a good promotion to the book, helping it to reach an even broader number of researchers.

Just one final remark: on the review there is an error on the link to the FLOWer database. The correct link is: http://flower.web.ua.pt/.

ISAC Congress - The plant sciences team

For prosperity the picture of the organizing team of the plant sciences sections of the ISAC Congress.

(from left to right: Jaroslav Dolezel, João Loureiro, David Galbraith, Jan Suda, Johann Greilhuber, Brian Husband and Paul Kron)

ISAC Congress - Day 5

And so the last day of the XXIV ISAC Congress came, and since the morning we could see the difference in the number of participants that were present in the Budapest Sportsarena. After 4 long days, the exhaustion was already felt, personally speaking, but there was still time for some last chatting with the commercial exhibiters and most importantly, to profit from another interesting workshop dedicated to the plant sciences. Jan Suda, Brian Husband and Paul Kron brought for the first time the world of plant evolutionary biology, biosystematics and ecology to the ISAC Congress, and I was very happy to witness that at least a dozen participants were interested. As expected, the workshop was very good with the speakers presenting several examples on the potentialities that flow cytometry has on such fields together with some "controversy" related topics on best practices.

Then was time to relax, drink some hungarian beers in a local pub and eat some typical food, before going to rest with the idea that everything was done to fulfil the objectives of the Congress. Of course, we leave the evaluation for the participants... and we hope that they have enjoyed as much as I did.

ISAC Congress - Day 4

The Day 4 of the now International Society of Advancements on Cytometry saw the comeback of the plant sciences, first by the presence of Jaroslav Dolezel in the Parallel Session of "High Throughput, High Content Analysis" with the very interesting talk on "Development of Flow-Based Strategies for Genomics of Crop Plants". There were also some interesting talks on the Advances in Flow Cytometry Instrumentation, especially those related with Acoustically Focused Sample Streams.

The day continued with the presentation of the FlowJo Software, this time dedicated to the bare basics. It seems quite a powerful tool, but still I would like to test it myself to evaluate which features may be of particular interest for plant DNA flow cytometry purposes. Again, and as with the new instruments, I will soon dedicate a post to the new software that was presented in the Congress.


In the afternoon the first Workshop dedicated to plants took place "Plant Genome Structure and Gene Expression", and to our surprise up to 25 people were present, which clearly surpassed our best expectations. The Workshop was wonderfully conducted by Jaroslav Dolezel and David Galbraith, and we had nice talks made by Jan Suda, Johann Greilhuber and by both chairmans, which covered several topics on the application of FCM in plant sciences, from sample preparation and storage, standards and standardizations, chromosome sorting for studies of genome structure and protoplasts and nuclei sorting for studies of gene expression. I had the opportunity to present some slides on the effect of cytosol on the quantitative staining of DNA, which despite being well received, w perhaps too technical for what was intended. However in each talk, some interesting questions were made and in the end we were satisfied with the overall output and reception of the workshop.

The poster session was just afterwards and again it was a nice opportunity to present the FLOWer database to many different people. The reception was good and it was a very nice idea of the congress organizers to put all the poster of similar areas close to each other.

By the end of the day, I was almost "dead" but there were still some forces for some good dinner and fruitful discussions.

Our poster on the FLOWer database presented in ISAC Congress 2008

The ISAC congress is coming...

The XXIV International Congress of ISAC is starting already in the following 17th of May and I look forward for the event. I will make a daily report of the congress, accompanied by some documenting pictures. So keep tuned...

For your appetizing I will leave you with the advertising that will be made of one of the highlights in what concerns plant flow cytometry - the Workshop "Plant Genome Structure and Gene Expression". I hope that all the people in the picture are heading to the Budapest Sportsarena for the workshop. We do not mind the sporty clothes!

Dear ISAC Congress Attendee:

Jaroslav Dolezel and David Galbraith would like to invite you to attend Workshop 5 “Plant Genome Structure and Gene Expression” at the International ISAC Congress in Budapest. This workshop is scheduled for 15:45-17:15 on Tuesday, 20 May 2008, and will be held in the Red Room.

This workshop is aimed not simply at scientists interested in employing flow and image cytometry for the manipulation of plant cells, tissues, organs, and organelles. It also will provide valuable information for Operators and Core Managers having no prior experience with plant samples, who increasingly will encounter plant scientists interested in using their technologies and instruments in their plant-based research.

Topics to be presented and discussed will include:

• Preparation and storage of samples for DNA flow cytometry.
• Combating the negative effect of cytosol on nuclear DNA staining.
• Standardization for reliable estimation of nuclear genome size.
• Chromosome isolation and sorting.
• Analysis of gene expression, including flow analysis and sorting of organelles and large cells.

There will also be opportunities provided for questions, comments, etc., from the audience.

WORKSHOP PROGRAM:

Moderators: Jaroslav Dolezel (Olomouc, CR) and David Galbraith (Tucson, USA)

Short presentations on each topic will be given by:

• Jan Suda (Charles University in Prague, Czech Republic)
• Johann Greilhuber (University of Vienna, Austria)
• João Loureiro (Charles University in Prague, Czech Republic)
• Jaroslav Dolezel (Institute of Experimental Botany, Olomouc, Czech Republic)
• David Galbraith (University of Arizona, Tucson, USA)

Open Discussion will follow.

ISAC XXIV International Congress, May 17-21, 2008, Budapest, Hungary
http://www.isac-net.org/congress2008/

Recent requests for help

I recently received two e-mails from some colleagues asking for some help in not so explored plant flow cytometry subjects.

You can see their messages below and if you have can give them any feedback to their question, they will certainly be very grateful.

E-mail 1:
"Dear all,

One groups wants to separate Lolium nuclei from the organelles... This project is really challenging for me and I'll need some help.

The main target is separating mitochondria (urgent application) and the second target is sorting chromosomes. Lolium has 7 chromosomes in the haploid set. As I told you I'm a little bit out of the Botany field now and I would like to come back... Do you have some protocols, procedures, papers, information, tips and tricks... that could help me? For analysis I think I shouldn't have big problems because of the equipments that I've in the lab, but for sorting I could be limited because I've a FACSAria and it has a 70 and a 100 um nozzles.

Nevertheless, probably the higher restriction for chromosome sorting it could be that instead of a UV laser the Aria has a violet laser...

Any help will be very welcome.

Kind regards,

Dr. Alfonso Blanco Fernández (e-mail: alfonso.blanco@ucd.ie)
Flow Cytometry Core Facilities
UCD - Conway Institute of Biomolecular & Biomedical Research
University College Dublin,
IRELAND"

E-mail 2:
"Hello Joao,

Please can I have a question – do you have an experience in plant cell sorting based on neutral lipids content – ideally using auto-fluorescence. We have a customer who is sorting algae and cyanobacteria based on neutral lipids content – and asked us for an advice in this..

Thanks,

Roman Navratil , Ph.D. (e-mail: Roman.Navratil@i-cyt.com)
Applications Specialist
iCyt Mission Technology, Inc.
Praha, CZECH REPUBLIC"

Your help is needed! You can reply to these problems either by commenting this post or by contacting directly the researchers through the e-mail given in each message.

The Annals of Botany Plant Genome Horizons Special Issue

As it was premiered in the blog 2 and 1/2 months ago, the Annals of Botany has just released a Special Issue dedicated to Plant Genomes in tribute of Michael Bennett's, a leading researcher on this area that decided to retire from the Jodrell Laboratory (Kew Royal Botanical Gardens, UK) in 2007.

As expected this is a very interesting issue of the Annals of Botany, that covers very different topics of plant genomes and opens future horizons. The issue is composed by a nice introduction that explores the contribution of Michael Bennett (in the picture) to Genome Research, some reviews and several research papers.

It is definitely worth reading.