Promoter analysis in transient assays using a GUS reporter gene construct in creeping bentgrass (Agrostis palustris).
Transient expression profiles for several chimeric beta-glucuronidase (GUS) gene constructs were determined in tissues (young leaves, mature leaves and roots) of creeping bentgrass (Agrostis palustris, cv. Penn A4) following microprojectile bombardment. The constructs analyzed consisted of the uidA (GUS) reporter gene driven by four different promoters (ubiquitin 3-potato, ubiquitin corn, ubiquitin rice and CaMV 35S). The total number of GUS hits (or transient expression units; TEUs) were determined manually under a dissecting scope after histochemical staining for GUS. Results suggest that the ubiquitin rice promoter is most active in cells of turfgrass, regardless of the developmental stage or tissue-type. The ubiquitin corn promoter was the next best. Of the four promoter used, except for ubiquitin 3-potato, reporter gene activity was dramatically higher in mature leaves compared to young leaves. The relative efficiency of each promoter was about the same in roots and leaves. We have also analyzed uidA (GUS) reporter gene activity following microprojectile bombardment in transient expression assays with callus from two cultivars (Providence or Penn A4) of creeping bentgrass. Differences in the frequency of GUS positive hits were observed between cultivars up to 72 hours post-bombardment. However, this difference between cultivars disappeared after 72 hours post-bombardment. This information describing promoter functionality in bentgrass will be important when designing gene constructs for trait modification and when choosing appropriate cultivars for improvement through gene transfer experiments. This is the first in depth report on organ-specific and developmental gene expression profiles for transgenes in a turfgrass species.
Basu
Chhandak
C
Department of Plant Sciences, University of Rhode Island, 9 East Alumni Avenue, Suite 7, Kingston, Rhode Island 02881, USA.
Kausch
Albert P
AP
Luo
Hong
H
Chandlee
Joel M
JM
D031703:Agrostis / Q000235:genetics*; D019470:Biolistics; D015870:Gene Expression; D017930:Genes, Reporter; D005966:Glucuronidase / Q000235:genetics; D018515:Plant Leaves / Q000235:genetics; D018517:Plant Roots / Q000235:genetics; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics
D011993:Recombinant Fusion Proteins; D005966:Glucuronidase