Specificity of expression of the GUS reporter gene (uidA) driven by the tobacco ASA2 promoter in soybean plants and tissue cultures.
Twelve independent lines were transformed by particle bombardment of soybean embryogenic suspension cultures with the tobacco anthranilate synthase (ASA2) promoter driving the uidA (beta-glucuronidase, GUS) reporter gene. ASA2 appears to be expressed in a tissue culture specific manner in tobacco (Song H-S, Brotherton JE, Gonzales RA, Widholm JM. Tissue culture specific expression of a naturally occurring tobacco feedback-insensitive anthranilate synthase. Plant Physiol 1998;117:533-43). The transgenic lines also contained the hygromycin phosphotransferase (hpt) gene and were selected using hygromycin. All the selected cultures or the embryos that were induced from these cultures expressed GUS measured histochemically. However, no histochemical GUS expression could be found in leaves, stems, roots, pods and root nodules of the plants formed from the embryos and their progeny. Pollen from some of the plants and immature and mature seeds and embryogenic cultures initiated from immature cotyledons did show GUS activity. Quantitative 4-methylumbelliferyl-glucuronide (MUG) assays of the GUS activity in various tissues showed that all with observable histochemical GUS activity contained easily measurable activities and leaves and stems that showed no observable histochemical GUS staining did contain very low but measurable MUG activity above that of the untransformed control but orders of magnitude lower than the constitutive 35S-uidA controls used. Low but clearly above background levels of boiling sensitive GUS activity could be observed in the untransformed control immature seeds and embryogenic cultures using the MUG assay. Thus in soybean the ASA2 promoter drives readily observable GUS expression in tissue cultures, pollen and seeds, with only extremely low levels seen in vegetative tissues of the plants. The ASA2 driven expression seen in mature seed was, however, much lower than that seen with the constitutive 35S promoter; less than 2% in seed coats and less than 0.13% in cotyledons and embryo axes. The predominate tissue culture specific expression pattern of the ASA2 promoter may be useful for genetic transformation of crops.
Inaba
Yoshimi
Y
Department of Crop Sciences, Edward R. Madigan Laboratory, University of Illinois, 1201 W. Gregory Drive, Urbana, IL 61801, USA.
Zhong
Wei Qun
WQ
Zhang
Xing-Hai
XH
Widholm
Jack M
JM
D000878:Anthranilate Synthase / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D035264:Flowers / Q000378:metabolism; D017930:Genes, Reporter; D005966:Glucuronidase / Q000032:analysis*; D018515:Plant Leaves / Q000378:metabolism; D018547:Plant Stems / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000196:embryology / Q000378:metabolism*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000032:analysis; D012639:Seeds / Q000378:metabolism; D013025:Glycine max / Q000196:embryology / Q000235:genetics*; D046509:Tissue Culture Techniques; D014026:Nicotiana / Q000235:genetics*; D014170:Transformation, Genetic
D011993:Recombinant Fusion Proteins; D005966:Glucuronidase; D000878:Anthranilate Synthase