Publication Date:
2002-07-06
Description:
The zebrafish has become a favorite organism for genetic analysis of vertebrate development, but methods for generating mutants by reverse genetic approaches have been lacking. We report a method to obtain stable mutants of a gene based on knowledge of the gene sequence only. Parental fish were mutagenized with N-ethyl-N-nitrosourea; in 2679 F1 fish, the rag1 gene was analyzed for heterozygous mutations by resequencing. In total, we found 15 mutations: 9 resulted in amino acid substitutions and 1 resulted in a premature stop codon. This truncation mutant was found to be homozygous viable and defective in V(D)J joining. Although presumably immune deficient, these homozygous rag1 mutant fish are able to reach adulthood and are fertile. As sperm samples from all 2679 F1 fish were collected and cryopreserved, we have in principle generated a mutant library from which mutants of most zebrafish genes can be isolated.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Wienholds, Erno -- Schulte-Merker, Stefan -- Walderich, Brigitte -- Plasterk, Ronald H A -- New York, N.Y. -- Science. 2002 Jul 5;297(5578):99-102.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Hubrecht Laboratory, Center for Biomedical Genetics, Uppsalalaan 8, 3584 CT, Utrecht, Netherlands.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/12098699" target="_blank"〉PubMed〈/a〉
Keywords:
Amino Acid Substitution
;
Animals
;
Codon, Terminator
;
Ethylnitrosourea
;
Female
;
Gene Library
;
Gene Rearrangement
;
Genes, Immunoglobulin
;
*Genes, RAG-1
;
Haplotypes
;
Heterozygote
;
Homeodomain Proteins/chemistry/genetics
;
Immunoglobulin Heavy Chains/genetics
;
Introns
;
Male
;
Mutagenesis
;
*Mutation
;
Mutation, Missense
;
Polymerase Chain Reaction
;
Polymorphism, Single Nucleotide
;
Recombination, Genetic
;
Zebrafish/*genetics/immunology/physiology
Print ISSN:
0036-8075
Electronic ISSN:
1095-9203
Topics:
Biology
,
Chemistry and Pharmacology
,
Computer Science
,
Medicine
,
Natural Sciences in General
,
Physics
Permalink
