Development and characterization of a generalized gene tagging system for higher plants using an engineered maize transposon Ac.
This report describes a series of transposon tagging vectors for dicotyledonous plants based on the maize transposable element Ac. This binary system includes the transposase (Ts) and the tagging element (Ds) on separate T-DNA vectors. Ts elements include versions in which transcription is driven either by the endogenous Ac promoter or by the cauliflower mosaic virus (CaMV) 35S promoter. Ds tagging element includes a gene conferring methotrexate (Mtx) resistance for selection and a supF gene to facilitate cloning of tagged sequences. The Ds element is flanked by a CaMV 35S promoter and the beta-glucuronidase (GUS) coding sequence so that GUS expression occurs upon excision of the element. We have transformed these Ts and Ds elements into tobacco and demonstrated that the Ts is functional with either promoter, and that the artificial Ds elements are capable of transposition. The amount of excision was found to depend upon both the individual Ts and Ds primary transformants used. Somatic excision of Ds was seen in up to 100% of progeny seedlings containing Ts and Ds. Germinal excision was detected in up to 48% of the progeny of plants containing both elements. Hence, this system can generate a sufficient number of events to be useful in gene tagging.
Fitzmaurice
W P
WP
Department of Botany, North Carolina State University, Raleigh 27695.
Lehman
L J
LJ
Nguyen
L V
LV
Thompson
W F
WF
Wernsman
E A
EA
Conkling
M A
MA
D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular / Q000379:methods; D003433:Crosses, Genetic; D004247:DNA; D004251:DNA Transposable Elements; D004351:Drug Resistance / Q000235:genetics; D005818:Genetic Engineering; D005819:Genetic Markers; D005966:Glucuronidase / Q000235:genetics; D008727:Methotrexate / Q000494:pharmacology; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000235:genetics / Q000378:metabolism; D010944:Plants / Q000235:genetics*; D010947:Plants, Toxic; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012231:Rhizobium; D012639:Seeds / Q000235:genetics; D014026:Nicotiana; D014170:Transformation, Genetic; D019895:Transposases; D003313:Zea mays / Q000235:genetics*
D004251:DNA Transposable Elements; D005819:Genetic Markers; D004247:DNA; D009713:Nucleotidyltransferases; D019895:Transposases; D005966:Glucuronidase; D008727:Methotrexate