23.10.2007

We have now uploaded 22.672 EST's to the GenBank. Out of a total number of 29.018 sequences, we were left with 22.672 after removing mitochindrial sequences, vector sequences and a general quality check. Please find more inforamtion regarding the different cDNA libraries below:

Name: Gadus morhua pooled liver samples SSH library (exposed to toxicants) ZNL
Total ESTs: 2304
Passed: 2004 (86%)
Average length:613

Name: Gadus morhua stem cell library ZNS
Total ESTs: 3935
Passed: 2719 (69%)
Average length:442

Name: Gadus morhua pooled liver samples library (exposed to toxicants) GML
Total ESTs: 4608
Passed: 4002 (86%)
Average length:593

Name: Gadus morhua intestinal library ZNC
Total ESTs: 3456
Passed: 3153 (91%)
Average length:563

Name: Gadus morhua pooled liver samples library KL
Total ESTs: 426
Passed: 410 (96%)
Average length:512

Name: Gadus morhua pooled kidney samples library ZNKAA
Total ESTs: 7353
Passed: 5408 (73%)
Average length:492

Name: N/A
Total ESTs: 10
Passed: 4 (40%)
Average length:621

Name: Gadus morhua pooled gill samples library ZNGAA
Total ESTs: 6879
Passed: 4924 (71%)
Average length:481

Name: Gadus morhua liver and intestine library TSTf
Total ESTs: 56
Passed: 47 (83%)
Average length:461

06.09.2006

Please find a list of the genes we have included on our small-scale Atlantic cod custom made cDNA array below. By clicking on the link you will download an excel spreadsheet displaying the genes on the array. The first-generation array will be printet in September 2006, and tested before applied in our ongoing research. The array contains some 450 stress-responsive genes, about 90 immuno-genes, about 80 metabolism genes, in addition to about 70 control or reference genes, all together about 750 clones. Some of the clones are encoding the same proteins, making the total number of genes on the array somewhat lower.

Atlantic cod cDNA array - genes

28.03.2006

A pipeline for automatic EST annotation has been established in collaboration with CBU. It's based on a triplicate BLAST system; BLASTX (protein) - BLASTN (nucleotide) - BLASTN (EST). Both best hit and best annotated hit are reported for protein and nucleotide BLAST's. That way we don't have to screen through hundreds of un-annotated Tetraodon and Danio sequences uploaded to the Genbank. Our database has also been made searchable, in that we can run BLASTN against our own sequences. These include about 15000 EST's, that consist of 1300 contigs and about 10000 singlets (representing 11300 genuine gene sequences). Unfortunately, the database will not be made public in so far. But if you are interested in certain genes, I can search for them within the database for you. And provide them upon request (and within an agreement on the use).

The pipeline was presented as a poster at the recent Havbruk 2006 conference in Bergen 29-31 March 2006.

14.2.2006

We have now assembled our 15.000 cod EST's with the automated annotation pipeline developed by the Computational Biology Unit (CBU) at the University of Bergen. A lot of interesting genes have been found. Soon we will start uploading these gene sequences to the GenBank. We have also started the process of selecting the EST's to be included on our custom-made array.

5.1.2006

We have now sequenced about 10000 EST clones from two cDNA libraries, one from gill tissue and one from kidney tissue. The sequences will be uploaded to the GenBank as soon as possible. They will also be used in assembly with the other 5000 EST sequences we have, in search for stress-related genes, to be used in array production.

16.12.2005

Årsrapport til NFR medio oktober 2005

Stress genes in the Atlantic cod Gadus morhua: EST sequencing and microarray production

Statusrapport for prosjektet "Stress genes in Atlantic cod Gadus morhua: EST sequencing and microarray production" (NFR prosjekt nr: 159197/I20). Juvenil torsk har blitt eksponert for ulike miljøgifter som PCB, PAH, bisfenol, bromerte flammehemmere, kopper og kadmium og RNA ekstrahert fra opp til 10 forskjellige vevstyper. I tillegg har RNA til EST sekvensering blitt samlet fra torsk som har blitt eksponert for vegetabilske fôrkilder. Deler av dette materialet har blitt brukt til å lage cDNA biblioteker. Tre cDNA biblioteker har så langt blitt konstruert fra vev fra torsk eksponert gjennom tvangsfôring for PCB, PAH, Cu og Cd, to biblioteker fra lever og et fra lever + tarm. 110 EST'er ble sekvensert fra det første lever + tarm cDNA biblioteket, men kvaliteten på biblioteket var ikke høy nok til å starte storskala shotgun EST-sekvensering. De gode EST sekvensene fra dette cDNA biblioteket vil bli lastet opp i Genbanken. Også det andre biblioteket av lever ble funnet å ha for dårlig kvalitet (lav titer) til massiv EST sekvensering. I det tredje cDNA biblioteket har vi sekvensert 24 kloner for kvalitetssjekk, og har nå bestilt 4600 EST kloner fra Amplicon Express (USA). I tillegg har det blitt sendt prøver til BGI LifeTech (Kina) for å få sekvensert ca. 5000 EST kloner fra et gjelle cDNA bibliotek og et tilsvarende antall kloner fra et nyre cDNA bibliotek. Real-time qRT-PCR protokoller har blitt etablert for en rekke stressgener i torsk, bl.a. antioksidant enzymer som superoxide dismutase, catalase og glutathione peroxidase, for heat shock protein (HSP) 70 og HSP90, CYP1A, metallothionein, ubiquitin, p53, pluss flere referansegener.

24.11.2005

We have now finally received 4600 EST clones from Amplicon Express, and are currently working with the nucleotide sequences. These EST's were picked from a liver cDNA library from cod force fed a mixture of toxicants (PCB, PAH, Cu and Cd). We are establishing a pipeline for annotation together with the Computational Biology Unit at the University of Bergen. In a couple of weeks all sequences will be clustered, and consensus sequences annotated. The results will made available for the global community as soon as they are ready. CBU are working with a database (OpenSputnik) that will be made available for the public. We also hope to upload these sequences to the GeneBank within a couple of months.

A company in China have recently made two cDNA libraries for us, one from gill tissue and one from head kidney tissue, and will sequence 5000 EST's from each library. I expect to receive these clones and EST sequences in December 2005, and we will also make them public as soon as possible. We have also, in collaboration with the Institute of Marine Research in Bergen, made another cDNA library from liver tissue from cod exposed to environmental pollutants, and sequenced the first 96 clones (one plate) with success last week. We will sequence at least 5 more plates from this library in the next weeks.

We will make a subtractive cDNA library from exposed cod the upcoming weeks, and search for weakly expressed and "on-off" genes in liver tissue.

A array with 2-300 oligos is planned made in the first half on 2006. This array will be spotted mainly by "stress genes", and used in various ongoing research projects.

The Institute of Marine Research in Bergen plans to make a first-generation cDNA array in early 2006, probably with clones from our liver stress cDNA library, but will also include clones from our gill cDNA library which we currently are sequencing in China. This array will be tested in winter 2006. It will contain about 20.000 gene sequences.

The National Research Council of Canada's Institute for Marine Biosciences in Halifax, Canada, recently received a grant of 15 mill. Canadian dollars for a genomic project on the Atlantic cod, and plans to do a massive sequencing effort. With the combined effort of Canadian, Icelandic and Norwegian research, tens of thousands of genes will most likely be sequenced in the coming months.

1.6.2005

A PhD student has been appointed to work with a cod microarray in the project "Effects of environmental and nutritional stress on gene expression in the Atlantic cod, Gadus morhua", which runs in parallel with this project.

01.07. 2005.

We are heavyly delayed in this project, partly due to the failure of the company Amplicon Express, USA, to sequence a cDNA library for cod liver.

20.05.2005

On a meeting on Iceland in May, it was decided to establish the International Cod Genomic Consortium. The preliminary web page of the consortium can be found here: http://www.icgcnet.info. The invitation to join the ICGC follows (written by Laura Brown, IRC of Canada):

Invitation to join the International Cod Genomics Consortium

Because of your expertise and experience in cod research and/or culture, we invite you to join the newly formed International Cod Genomics Consortium (ICGC).

Background: Dr. Ole Torrissen, Director of the Marine Research Institute in Bergen Norway, Dr. Sjöfn Sigurisladóttir, General Director of the Iceland Fisheries Laboratory in Reykjavik, Iceland, and Dr. Laura Brown, Senior Research Officer, National Research Council of Canada's Institute for Marine Biosciences in Halifax, Canada, engaged in several discussions on proposed and current cod genomics research within their respective countries.

It was clear that the approaches taken by each country and the research and technological strengths were similar or at least complementary. Therefore it seemed logical to combine resources among the three countries, and discuss the possibilities of sharing information, data, and resources.

Ole, Sjöfn, and Laura organized an informal workshop to discuss their research, share ideas, and to identify areas of research where collaborative efforts would benefit all. The workshop was held in Reykjavik, Iceland, May 10-11, 2005. Participants were to leave the meeting with the following questions answered or with a plan on how to answer these questions:

1. What cod research is being conducted in each country, and by whom?
2. What are one or two specific research programs we can work together?
1. These may include sequencing the entire genome
1. What information, data, resources and material can we share?
2. How can we pay for these collaborative programs?
3. How will this international collaborative venture be structured, i.e., who is leading it and who is responsible for keeping us on track?

The three organizers wanted to maintain the informality of this workshop, in order that participants would feel comfortable in sharing information and bringing out ideas of how to develop an international cod genomics consortium.

Outcomes of the Workshop: All participants shared information freely, discussions were open and frank, and there was a general consensus that most of the challenges for the cod farming industry and the cod fisheries were the same in all three countries.

Some of the most pressing biological challenges were identified as:

* Early maturation
* Larval qualtiy and juvenile deformities
* Vaccines/immunity
* Wild/farmed interactions
* Markers for growth, fecundity, quality
* Tools to examine early nutrition and health
* Alternative feeds for grow-out
* Welfare issues and ethics with new technologies

Other issues that were raised included:

* How can researchers interact with industry - how can industry quickly use the results of research?
* IP issues
* Data release policies
* How can our existing programmes interact?
* Should we invited others?
* Shoud we sequence the entire genome?

Decisions: It was decided that we would form the International Cod Genomics Consortium, with the following draft mandate:

The Internation Cod Genomics Consortium (ICGC) is a group of researchers from Iceland, Norway and Canada, and other countries, who share a common interest in research on cod (Gadus morhua), using genomics and other biotechnologies. The ICGC will work together to develop a plan to sequence the entire genome of the cod. The ICGC will also develop processes and media whereby they can share data, information, resources and technology, so that our knowledge of the biology and genomics of the cod in order to help strengthen a growing and environmentally sustainable aquaculture industry and fishery. Specific areas of biological interest are: health, nutrition, reproductive fitness, broodstock markers and environmental sustainability. Genomics technologies that are in common include EST libraries, gene sequencing, proteomics, cDNA microarrays, cloning, cell culture. The ICGC will develop databases where information may be posted and downloaded. Other bioinformatics solutions will be developed. The ICGC will hold a workshop once a year to continue the information and network development process.

Next steps:

1. Participants of the workshop will be asked who else should be invited to join the ICGC.
2. IFL will develop a webpage where documents can be posted and downloaded by the ICGC members
3. ICGC members will be sent a questionnaire to gather more information about existing projects, major technologies used, libraries contstructed, etc.
4. A summary will be sent to all members.
5. A database will be developed to deposit sequences, clone information, challenge experiments, etc.
6. Funding and development of a plan for genome sequencing will be formulated.
7. A workshop for 2006 will be planned - probably in Halifax.

If you are interested in joining the ICGC, please e-mail Dr. Þorleifur Ágústsson at tolli@rf.is with information about your research, current and proposed, approaches and technologies used, and your contact information. Currently there is no membership fee.

01.10.2005

4600 EST's will be sequenced from a cod liver library made from fish exposed to a mixture of toxicants. The sequencing will be fulfilled in November 2005, and the task has been outsourced to the american company Amplicon Express.

1.11.2005

10.000 EST's will be sequenced from two cDNA libraries from gill and head kidney tissues. The sequencing will be done by BGI LifeTech, Beijing Genomics Institute, in China. We will have the clones and the sequences as of December 2005. All together, the approximately 15.000 EST sequences will be used to design a custom-made microarray. This array should be finished during the winter of 2006, and will be used in ongoing research.