The movements of BK(Ca) channels were investigated in live cells using quantum dots (QDs). The extracellular N-terminus was metabolically tagged with biotin, labeled with streptavidin-conjugated QDs and then monitored using real-time time-lapse imaging in COS-7 cells and cultured neurons. By tracking hundreds of channels, we were able to determine the characteristics of channel movements quantitatively. Channels in COS-7 cells exhibited a confined diffusion in an area of 1.915 μm(2), with an initial diffusion coefficient of 0.033 μm(2)/s. In neurons, the channel movements were more heterogeneous and highly dependent on subcellular location. While the channels in soma diffused slowly without clear confinement, axodendritic channels showed more rapid and pseudo-one-dimensional movements. Intriguingly, the channel movement in somata was drastically increased by the neuronal β4 subunit, in contrast to the channels in the axodendritic area where the mobility were significantly decreased. Thus, our results demonstrate that the membrane mobility of BK(Ca) channels can be greatly influenced by the expression system used, subunit composition, and subcellular location. This QD-based, single-molecule tracking technique can be utilized to investigate the cellular mechanisms that determine the mobility as well as the localization of various membrane proteins in live cells.

We propose using O-desmethylvenlafaxine/venlafaxine for CYP2D6 phenotyping, and O-desmethylvenlafaxine/venlafaxine with venlafaxine + O-desmethylvenlafaxine for predicting venlafaxine treatment outcomes in future prospective studies.

Calcium-dependent gating of large-conductance calcium-activated potassium (BK(Ca)) channels is mediated by the intracellular carboxyl terminus, which contains two domains of regulator of K(+) conductance (RCK). In mammalian BK(Ca) channels, the two RCK domains are separated by a protein segment of 101 residues that is poorly conserved in evolution and predicted to have no regular secondary structures. We investigated the functional importance of this loop using a series of deletion mutations. We found that the length, rather than the specific sequence at the central region of the segment, is critical for the functionality of the channel. As the length of the loop is progressively shorted, the conductance-voltage relationship gradually shifts toward more positive voltages with a minimum length of 70 amino acids, in an apparent response to increased tension within the loop. Thus, the functional activity of the BK(Ca) channel can be modulated by altering the tension of this loop region.

Four-leaf white clover is not found easily in nature due to its low appearance rate (1 in 10,000). Because people believe that it brings good luck and like to either keep it or present it to a loved one, it has commercial and ornamental value. To breed four-leaf clover, we exposed its flowers to c-rays at the pollination stage. The M 1 seeds produced following doses at 25-100 Gy showed an approximately 74% germination rate, with seedling survival at 46%. In the M 1 generation of plants irradiated within that dose range, we found an increased frequency of four leaflets. One of them, Jeju Lucky-1 (JL-1), had a frequency of about 60%. To see whether that mutation was somaclonal or genetic, we observed its M 2 generation and found that such a phenotype reappeared. Although our results demonstrated that the irradiation of fully mature flowers led to a higher frequency of 4-leaflets, we could not clearly explain the genetic mechanism involved. We suggest that JL-1 is valuable as a new variety, without further genetic fixation, because white clover can be propagated vegetatively by stolons.

Multi point fluorescence measurement system using basic image shifting method and commercial multi mode fiber is presented in this paper. Using a singlet lens, the original fluorescence image of a sample is shifted to another plane which can be monitored using ccd, and at the first image plane independent two fiber tips in an xyz stage deliver each fluorescent signal at a specific sample position to a fluorescence correlation spectroscopy (FCS) with an electron multiplying charge coupled device (EMCCD). The FCS is composed with an EMCCD, which can detect single molecule level fluorescence light. Applying region of interest (ROI) and pixel binning, a time resolution of up to 2 ms can be achieved, which is sufficient to resolve the diffusion of fluorescence micro-sphere in solution. The advantages of implementing EMCCD cameras in wide-field ultra low light imaging, as well as in site-specific multi-point fluorescence measurement system, can consequently also be exploited for spatially and spectrally resolved FCS. Experimental results about FCS with spectrum informations demonstrate the advantage of the simplicity and flexibility of our system. We expect that this multi point measurement system also can be applied to other study of bio molecular dynamics.