Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Xinhong Chen
Date created: 2015-09-13 11:00:00
Date modified: 2015-09-15 09:55:48
pmyo3-ChR2-YFP ( C.elegans )
| Types | DnaRegion |
| Roles | engineered_region Composite |
| Sequences | BBa_K1634004_sequence (Version 1) |
Description
???myo-3 encodes MHC A, the minor isoform of MHC (myosin heavy chain) that is essential for thick filament formation, and for viability, movement, and embryonic elongation; expressed in body muscle, the somatic sheath cell covering the hermaphrodite gonad, and also expressed in enteric muscle, vulval muscles of the hermaphrodite and the diagonal muscles of the male tail. ???(from Wormatlas) Since we want to control the C.elegans' movement.We decided to use pmyo-3 (promoter of myo-3) to construct a plasmid which can drive the expression in the worm???s body muscle.ChR2 is a kind of channelrhodopsin. Channelrhodopsins are a subfamily of retinylidene proteins (rhodopsins) that function as light-gated ion channels.[1] They serve as sensory photoreceptors in unicellular green algae, controlling phototaxis: movement in response to light.[2] When we express the channelrhodopsins in some specific cells in organisms and shed specific light on them, we can activate or supress the specific ion channel to change the activity of the cell. According to this thought in optogenetic, we linked the Pmyo2，specific promoter in the muscle of head, with the ChR2, then we use the light source assembled by ourselves to shed light of specific wave on the C.elegans,changging the condition of the muscle. Since we have learned that the chosing of direction in C.elegans depends on the muscle in the head, we can observe the obvious change in moving pattern of the C.elegans after we shed the light. In other words,we try to construct a light-sensed locomotion controlling system in C.elegans. And we achieve that goal by using this part to control the movement of the head, the movement of the whole C.elegans.References:
1. Nagel G, Ollig D, Fuhrmann M, Kateriya S, Musti AM, Bamberg E, Hegemann P (June 2002). "Channelrhodopsin-1: a light-gated proton channel in green algae". Science 296 (5577): 2395???8. doi:10.1126/science.1072068
2. Sineshchekov OA, Jung KH, Spudich JL (June 2002). "Two rhodopsins mediate phototaxis to low- and high-intensity light in Chlamydomonas reinhardtii". Proc. Natl. Acad. Sci. U.S.A. 99 (13): 8689???94. doi:10.1073/pnas.122243399.
Notes
Since we use the restriction enzyme site to do the ligation, we should pay attention to the sequence of Pmyo3 and ChR2-YFP, avoiding using the restriction site exits in it.Source
We get Pmyo3 from the C.elegans' genomic DNA by PCR. ChR2 is a channelrhodopsin in green algae, but we didn't get it directly from the algae. Instead, we get ChR2 from some plamids by PCR. Then we ligated the Pmyo3 into PPD95.75, a vector commonly used in C.elegans. Finally, we construted the fused protein, ChR2-YFP,and ligated it into PPD95.75_Pmyo3.| Access | Instance | Definition |
| public public | BBa_K1634005 BBa_K1634003 | BBa_K1634005 BBa_K1634003 |
| Sequence Annotation | Location | Component / Role(s) |
| BBa_K1634005 BBa_K1634003 | 1,814 821,2629 | BBa_K1634005 BBa_K1634003 |