A microassay for quantitative determination of β-glucuronidase reporter gene activities in individually selected single higher plant cells
作者： B. Spörlein , A. Mayer , G. Dahlfeld , H.U. Koop
DOI: 10.1016/0168-9452(91)90163-3
关键词:
摘要: Abstract Using computer controlled microtechniques for individually selecting specific single cells and delivering maintaining volumes of liquids in the nanoliter range we have developed a procedure to quantitatively assay β-glucoronidase (GUS) activity individual transformed cells. standard solutions reaction product 4-methylumbelliferone (MU) determining appropriate optical parameters microscope photometer established suitability method quantitative determinations at cell level. The enzyme was performed after lysis subsequent incubation total volume 100 nl buffer, detection activities derived from easily achieved with 4-methylumbelliferyl glucuronide (MUG) as substrate. used address following problems: (1) variability GUS-activity among different tobacco mesophyll protoplasts stably plants, (2) determination efficiency promoters regulating same reporter gene level, (3) proportion transiently expressing GUS-gene PEG treatments (4) transient expression intranuclear microinjection DNA.
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