Reproducibility of frequency-dependent low frequency fluctuations in reaction time over time and across tasks.

OBJECTIVE: Increased levels of reaction time variability (RTV) are characteristics of sustained attention deficits. The clinical significance of RTV has been widely recognized. However, the reliability of RTV measurements has not been widely studied. The present study aimed to assess the test-retest reliability of RTV conventional measurements, e.g., the standard deviation (SD), the coefficient of variation (CV), and a new measurement called the amplitude of low frequency fluctuation (ALFF) of RT. In addition, we aimed to assess differences and similarities of these measurements between different tasks. METHOD: Thirty-seven healthy college students participated in 2 tasks, i.e., an Eriksen flanker task (EFT) and a simple reaction task (SRT), twice over a mean interval of 56 days. Conventional measurements of RTV including RT-SD and RT-CV were assessed first. Then the RT time series were converted into frequency domains, and RT-ALFF was further calculated for the whole frequency band (0.0023-0.167 Hz) and for a few sub-frequency bands including Slow-6 (<0.01 Hz), Slow-5 (0.01-0.027 Hz), Slow-4 (0.027-0.073 Hz), and Slow-3 (0.073-0.167 Hz). The test-retest reliability of these measurements was evaluated through intra-class correlation (ICC) tests. Differences and correlations between each EFT and SRT measurement were further examined during both visits. RESULTS: 1) The RT-ALFF of the Slow-5/4/3 and conventional measurements of RT-SD and RT-CV showed moderate to high levels of test-retest reliability. EFT RT-ALFF patterns generated slightly higher ICC values than SRT values in higher frequency bands (Slow-3), but SRT RT-ALFF values showed slightly higher ICC values than EFT values in lower frequency bands (Slow-5 and Slow-4). 2) RT-ALFF magnitudes in each sub-frequency band were greater for the SRT than those for the EFT. 3) The RT-ALFF in the Slow-4 of the EFT was found to be correlated with the RT-ALFF in the Slow-5 of the SRT for both two visits, but no consistently significant correlation was found between the same frequency bands. CONCLUSIONS: These findings reveal good test-retest reliability for conventional measurements and for the RT-ALFF of RTV. The RT-ALFF presented frequency-dependent similarities across tasks. All of our results reveal the presence of different frequency structures between the two tasks, and thus the frequency-dependent characteristics of different tasks deserve more attention in future studies.

[Heterogenous expression of nitrile hydratase gene of Nocardia sp. in recombinant Pichia pastoris].

Heterogenous expression of active nitrile hydratase (NHase) was focused for its great potential in genetically evolution of the operational stability of NHase. Two recombinant Escherichia coli strains, E. coli JM105 (pUC18-NHBAX) and E. coli BL21 (DE3) (pET32a-NHBAX), were first constructed and used for heterogenous expression of a NHase gene cloned from Nocardia YS-2002. It was found that the alpha subunit of NHase can not be effectively expressed in both recombinant E. coli, which results in as low as 0.04U/mg (dry cell weight) activity of NHase in E. coli BL21 (DE3) (pET32a-NHBAX), and no activity in JM105 (pUC18-NHBAX) at all. Therefore, the recombinant and active expression of NHase in Pichia pastoris was especially interested. A new plasmid, pPIC3.5k-NHBAX, was constructed by insertion of the NHase gene, and then successfully transformed into the cell of Pichia pastoris GS115 by electroporation. A novel superior strain, P. pastoris NH4, was selected from 6 target-clones and used for optimization of cell culture and NHase expression. The final activity of NHase expressed in P. pastoris NH4 under optimal conditions is 0.52U/mg (dry cell weight), which is 13-fold higher than that in recombinant E. coli. However, the activity of NHase expressed in recombinant P. pastoris NH4 can not be stably maintained longer than 6 h.