Frequently Asked Questions (FAQ)
How are ZeneMark® zebrafish lines used for research that ultimately benefits human health?
What is the nature of the retroviral insertions in ZeneMark® library fish?
How do I identify my favorite gene in Zv7 and pick an insertion?
How do I initiate a collaboration with Znomics in order to analyze mutations in my favorite genes?
Are insertions into introns or promoter regions mutagenic?
How do I genotype the ZeneMark® lines?
What is the genetic background of ZeneMark® lines?
Will more insertions be added to the database?
How are ZeneMark® zebrafish lines used for research that ultimately benefits human health?
Zebrafish are used by many laboratories around the world to do research on the basic molecular mechanisms of organ function and disease that are shared by both fish and humans. Over 1,000 research papers describing work on zebrafish are published each year in leading scientific journals. The international Zebrafish Model Organism Database is an excellent source of information about zebrafish research. There is also an ever-growing number of research papers and reviews available at the National Center for Biotechnology Information website. The ZeneMark Library contains targeted gene mutations for use by scientists worldwide to do research that is highly relevant to human disease.
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What is the nature of the retroviral insertions in ZeneMark® zebrafish?
The library was generated by insertion of a 7 kb retrovirus into zebrafish chromosomes using a highly mutagenic virus developed at Znomics. The LTR primer and target sequences are available to investigators who use ZeneMark lines in collaborative studies.
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How do I identify my favorite gene in Zv7 and pick an insertion?
Our posting of the new ZeneMark® Zv7 Map adds an important new resource for the zebrafish community. Znomics has mapped its entire insertion library to the Ensembl Zv7 edition of the zebrafish genome. The ZeneMark® Library web pages include a tutorial on how to search for your favorite gene using BLAST and how to pick insertions based on the Zv7 map. If you need additional assistance with selecting insertions, please contact us. We are happy to help.
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How do I initiate a collaboration with Znomics in order to analyze mutations in my favorite genes?
Use the convenient Research Inquiry form to provide us with your contact information and to describe the nature of the collaboration for which you wish to engage us and/or to specify the zebrafish lines in which you are interested. Or call 503-827-5271, ext. 201.
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What is the difference between a new ZeneMark® Library line and a Living Library line or a Human Disease Library line?
Living Library and Human Disease Library lines are subsets of the ZeneMark® Library. Original ZeneMark® Library lines and Human Disease Library lines are new lines that must be re-identified as described below. Living Library Lines are previously-identified zebrafish lines that are kept as live fish or frozen sperm samples.
The ZeneMark® Library consists of pools of frozen sperm obtained from retrovirus-injected fish. For any gene insertion requested, Znomics reconstitutes a line containing the specified insertion by first screening through the library pools by PCR using a primer specific to the virus and a primer specific to the flanking genomic sequence. Once the sperm sample containing the insertion is identified, Znomics uses that sperm to fertilize embryos in vitro to generate an F1 generation. These F1 progeny are raised at Znomics until they are of suitable age for tail biopsy and genotyping. The F1 zebrafish are genotyped at Znomics by PCR analysis on tail DNA in order to identify heterozygous carriers of the desired insertion. Insertion-positive fish are then out-crossed to a wild type strain.
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Are insertions into introns or promoter regions mutagenic?
Many intronic or promoter insertions are mutagenic, but we have no systematic way of determining mutagenicity for a specific insertion. Previous studies from other labs using retroviral mutagenesis methods have shown that the majority of mutations identified were due to intronic insertions.
For more information, please refer to:
Golling, G. et al, 2002. Insertional mutagenesis in zebrafish rapidly identifies genes essential for early vertebrate development.
Nat. Genet. 2002 Jun;31(2):135-40
Amsterdam, A. 2003. Insertional mutagenesis in zebrafish.
Dev. Dyn. 2003 Nov;228(3):523-34
Amsterdam, A., et al, 2004. Identification of 315 genes essential for early zebrafish development.
Proc. Natl. Acad. Sci. U S A. 2004 Aug 31;101(35):12792-7.
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How do I genotype the ZeneMark® lines?
We recommend standard DNA isolation and nested PCR methods. Primer sequences are provided to purchasers along with the DNA sequence of insertion sites. We use primer3, an open source program, to pick high quality primers. We are happy to consult if you experience difficulty with genotyping.
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What is the genetic background of ZeneMark lines?
The ZeneMark® Library was created on, and is out-crossed to, wild-type background zebrafish derived from EK and Florida strain wild-type zebrafish. This strain has high sequence similarity to the TU strain. Znomics wild-type zebrafish are available on request.
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Will more insertions be added to the database?
Yes. We are continuously updating the Znomics Libraries as new insertion junctions are sequenced and mapped. Investigators may sign up for our automated notification system to receive emails when a new insertion maps to their genes of interest.
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