The molecular deposition of transgenically modified starch in the starch granule as imaged by functional microscopy.
The molecular deposition of starch extracted from normal plants and transgenically modified potato lines was investigated using a combination of light microscopy, environmental scanning electron microscopy (ESEM) and confocal laser scanning microscopy (CLSM). ESEM permitted the detailed (10 nm) topographical analysis of starch granules in their hydrated state. CLSM could reveal internal molar deposition patterns of starch molecules. This was achieved by equimolar labelling of each starch molecule using the aminofluorophore 8-amino-1,3,6-pyrenetrisulfonic acid (APTS). Starch extracted from tubers with low amylose contents (suppressed granule bound starch synthase, GBSS) showed very little APTS fluorescence and starch granules with low molecular weight amylopectin and/or high amylose contents showed high fluorescence. Growth ring structures were sharper in granules with normal or high amylose contents. High amylose granules showed a relatively even distribution in fluorescence while normal and low amylose granules had an intense fluorescence in the hilum indicating a high concentration of amylose in the centre of the granule. Antisense of the starch phosphorylating enzyme (GWD) resulted in low molecular weight amylopectin and small fissures in the granules. Starch granules with suppressed starch branching enzyme (SBE) had severe cracks and rough surfaces. Relationships between starch molecular structure, nano-scale crystalline arrangements and topographical-morphological features were estimated and discussed.
Center for Molecular Plant Physiology, Plant Biochemistry Laboratory, Department of Plant Biology, The Royal Veterinary and Agricultural University, 40 Thorvaldsensvej, DK-1871 Frederiksberg C, Copenhagen, Denmark. abl@kvl.dk
Andreas
A
Blennow
Michael
M
Hansen
Alexander
A
Schulz
Kirsten
K
Jørgensen
Athene M
AM
Donald
James
J
Sanderson
D005456:Fluorescent Dyes; D007091:Image Processing, Computer-Assisted; D008853:Microscopy; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning; D010316:Particle Size; D030821:Plants, Genetically Modified; D011721:Pyrenes; D011198:Solanum tuberosum / Q000737:chemistry / Q000235:genetics; D013213:Starch / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure
C098210:1-aminopyrene-3,6,8-trisulfonic acid; D005456:Fluorescent Dyes; D011721:Pyrenes; D013213:Starch