PMC:4307275 / 25329-26604 JSONTXT

{"project":"2_test","denotations":[{"id":"25561231-11334735-50799913","span":{"begin":463,"end":465},"obj":"11334735"},{"id":"25561231-11401292-50799914","span":{"begin":892,"end":894},"obj":"11401292"},{"id":"25561231-21914652-50799915","span":{"begin":895,"end":897},"obj":"21914652"},{"id":"25561231-20038550-50799916","span":{"begin":898,"end":900},"obj":"20038550"},{"id":"T14406","span":{"begin":463,"end":465},"obj":"11334735"},{"id":"T30839","span":{"begin":892,"end":894},"obj":"11401292"},{"id":"T81653","span":{"begin":895,"end":897},"obj":"21914652"},{"id":"T56239","span":{"begin":898,"end":900},"obj":"20038550"}],"text":"Also, since our strategy of FCA using protoplasts is based on targeting fluorescent proteins into chloroplasts, the autofluorescence derived from chlorophylls of chloroplasts should be also considered. It has been reported that plant derived-autofluorescence causes some difficulties with some analytical processes of fluorescence-based techniques, and that chlorophylls of chloroplasts have been considered as one of the strongest autofluorescence contributors [26]. However, several previous reports have shown that only if the commonly used fluorescence markers such as eGFP, YFP and mCherry is used for the fluorescence-based techniques, the autofluorescent background from chlorophylls could be almost spectrally filtered out, because the autofluorescence background of chlorophylls is bathochromically shifted with respect to the fluorescent signals of eGFP, YFP, and mCherry proteins [27,28,29]. Since the fluorescent protein used in this study is a sGFP identical to commercially available eGFP (Clontech; TaKaRa Bio, Shiga, Japan), it could be supposed that the autofluorescence background of chlorophylls is not too much trouble with our strategy of chloroplast targeting-based FCA, because it can be almost filtered out, based on the bathochromic shift among them."}