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Proceedings of the National Academy of Sciences
Article . 1993 . Peer-reviewed
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Target-selected gene inactivation in Caenorhabditis elegans by using a frozen transposon insertion mutant bank.

Authors: R R, Zwaal; A, Broeks; J, van Meurs; J T, Groenen; R H, Plasterk;

Target-selected gene inactivation in Caenorhabditis elegans by using a frozen transposon insertion mutant bank.

Abstract

To understand how genotype determines the phenotype of the animal Caenorhabditis elegans, one ideally needs to know the complete sequence of the genome and the contribution of genes to phenotype, which requires an efficient strategy for reverse genetics. We here report that the Tc1 transposon induces frequent deletions of flanking DNA, apparently resulting from Tc1 excision followed by imprecise DNA repair. We use this to inactivate genes in two steps. (i) We established a frozen library of 5000 nematode lines mutagenized by Tc1 insertion, from which insertion mutants of genes of interest can be recovered. Their address within the library is determined by PCR. (ii) Animals are then screened, again by PCR, to detect derivatives in which Tc1 and 1000-2000 base pairs of flanking DNA are deleted, and thus a gene of interest is inactivated. We have thus far isolated Tc1 insertions in 16 different genes and obtained deletion derivatives of 6 of those.

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Keywords

Base Sequence, Nematoda, Molecular Sequence Data, DNA, Polymerase Chain Reaction, Mutagenesis, Insertional, Gene Expression Regulation, Oligodeoxyribonucleotides, Freezing, DNA Transposable Elements, Escherichia coli, Animals, Caenorhabditis elegans, Alleles, Gene Deletion, Gene Library

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    selected citations
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    269
    popularity
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    Top 10%
    influence
    This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
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    impulse
    This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
    Top 1%
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selected citations
These citations are derived from selected sources.
This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Citations provided by BIP!
popularity
This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.
BIP!Popularity provided by BIP!
influence
This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Influence provided by BIP!
impulse
This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
BIP!Impulse provided by BIP!
269
Top 10%
Top 1%
Top 1%
bronze