Developmentally regulated site-specific marker gene excision in transgenic B. napus plants.
We have developed a self-excision Cre-vector to remove marker genes from Brassica napus. In this vector cre recombinase gene and bar expression cassette were inserted between two lox sites in direct orientation. These lox-flanked sequences were placed between the seed-specific napin promoter and the gene of interest (vstI). Tissue-specific cre activation resulted in simultaneous excision of the recombinase and marker genes. The vector was introduced into B. napus by Agrobacterium-mediated transformation. F1 progeny of seven lines with single and multiple transgene insertions was subjected to segregation and molecular analysis. Marker-free plants could be detected and confirmed by PCR and Southern blot in all transgenic lines tested. The recombination efficiency expressed as a ratio of plants with complete gene excision to the total number of investigated plants varied from 13 to 81% dependent on the transgene copy number. Potential application of this system would be the establishment of marker-free transgenic plants in generatively propagated species.
Kopertekh
Lilya
L
Julius Kühn Institute, Federal Research Centre for Cultivated Plants (JKI), Institute for Biosafety of Genetically Modified Plants, Erwin-Baur-Str. 27, 06484, Quedlinburg, Germany.
Broer
Inge
I
Schiemann
Joachim
J
D029688:Brassica napus / Q000235:genetics*; D018744:DNA, Plant / Q000235:genetics; D004274:DNA, Recombinant / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D005819:Genetic Markers; D005822:Genetic Vectors; D019426:Integrases / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics; D017422:Sequence Analysis, DNA; D014169:Transformation, Bacterial; D019076:Transgenes
D018744:DNA, Plant; D004274:DNA, Recombinant; D005819:Genetic Markers; C045073:Cre recombinase; D019426:Integrases