High frequency of phenotypic deviations in Physcomitrella patens plants transformed with a gene-disruption library.
BACKGROUND
The moss Physcomitrella patens is an attractive model system for plant biology and functional genome analysis. It shares many biological features with higher plants but has the unique advantage of an efficient homologous recombination system for its nuclear DNA. This allows precise genetic manipulations and targeted knockouts to study gene function, an approach that due to the very low frequency of targeted recombination events is not routinely possible in any higher plant.


RESULTS
As an important prerequisite for a large-scale gene/function correlation study in this plant, we are establishing a collection of Physcomitrella patens transformants with insertion mutations in most expressed genes. A low-redundancy moss cDNA library was mutagenised in E. coli using a derivative of the transposon Tn1000. The resulting gene-disruption library was then used to transform Physcomitrella. Homologous recombination of the mutagenised cDNA with genomic coding sequences is expected to target insertion events preferentially to expressed genes. An immediate phenotypic analysis of transformants is made possible by the predominance of the haploid gametophytic state in the life cycle of the moss. Among the first 16,203 transformants analysed so far, we observed 2636 plants (= 16.2%) that differed from the wild-type in a variety of developmental, morphological and physiological characteristics.


CONCLUSIONS
The high proportion of phenotypic deviations and the wide range of abnormalities observed among the transformants suggests that mutagenesis by gene-disruption library transformation is a useful strategy to establish a highly diverse population of Physcomitrella patens mutants for functional genome analysis.
Plant Biotechnology, Freiburg University, Sonnenstrasse 5, D-79104 Freiburg/Br, Germany. tanja.egener@biologie.uni-freiburg.de
Tanja
T
Egener
José
J
Granado
Marie-Christine
MC
Guitton
Annette
A
Hohe
Hauke
H
Holtorf
Jan M
JM
Lucht
Stefan A
SA
Rensing
Katja
K
Schlink
Julia
J
Schulte
Gabriele
G
Schween
Susanne
S
Zimmermann
Elke
E
Duwenig
Bodo
B
Rak
Ralf
R
Reski
D019068:Bryopsida / Q000235:genetics* / Q000254:growth & development; D004251:DNA Transposable Elements; D018076:DNA, Complementary / Q000235:genetics; D015723:Gene Library; D018745:Genome, Plant; D016296:Mutagenesis; D016254:Mutagenesis, Insertional; D009154:Mutation; D010641:Phenotype; D030821:Plants, Genetically Modified; D011995:Recombination, Genetic; D014170:Transformation, Genetic
D004251:DNA Transposable Elements; D018076:DNA, Complementary