Root sampling method for aquaporin gene expression analysis in rice

A rice root system consists of main roots, and lateral roots which account for a large portion of the total length and surface area of the whole root system, indicating that lateral roots greatly contribute to its hydraulic conductance. It is therefore worthwhile measuring aquaporin activities of lateral roots in comparison with main roots to evaluate their contribution to hydraulic conduc-tivity. However, the sampling of fine lateral roots is time-consuming, and its method can be critical be-cause of the difficulties of handling them for sam-pling. In addition, root has been reported to be sensitive to the surrounding environment and af-fected by destructive sampling procedures in which lateral roots are removed from main roots. There-fore, in this study, we attempted to establish a method for collecting samples of lateral and main roots for aquaporin analysis while minimizing the effect of separation of roots from the shoot, and the removable of lateral roots from main roots on the transcript abundance. We conducted three ex-periments and measured the expression levels of four aquaporin genes including OsPIP2;1, Os-PIP2;4, OsPIP2;5 and OsTIP2;1 with different sampling time length (experiment 1), with or without shoot removal (experiment 2) and light and dark periods samplings with different light condi-tions (experiment 3). Our results revealed that the aquaporin expression levels in separated roots did not change compared to those of the roots which were immediately preserved after sampling from the shoot, when root separation was conducted within 10 minutes without shoot removal under flu-orescent light during light period of growth, as well as under indirect lighting during dark period of growth.