Information Technology Reference
In-Depth Information
“sender”
“receiver”
main
metabolism
TetO
TetR
LuxI
VAI
GFP
[aTc]
UV
Fluorescence
Figure 7.20
Circuit diagram of gradient communications.
ious columns were induced with different levels of aTc. Also, one control
column included receivers that were induced directly with the VAI extract.
Figure 7.21 shows the results of this experiment after culturing the plate for
4 h at 37°C. As expected, the null wells containing no aTc or VAI showed no
enhancement of fluorescence, while the positive control wells with the 10
×
VAI extract exhibited fluorescence. The experiments labeled BL21-LuxITet
include senders where the pLuxI-Tet-8 plasmid was transformed into BL21-
PRO cells, while the experiments labeled DH5
-LuxITet include senders
where the pLuxI-Tet-8 plasmid was transformed into
E. coli
DH5
α
cells.
1
In wells containing sender cells induced with aTc at levels lower than about
20 ng/ml, the receiver cells exhibit only only a small fluorescent response.
In wells induced with aTc levels great than 200 ng/ml, the receiver cells
exhibit a significant response. Sufficiently high levels of aTc inhibited cell
growth.
Visual Observation of Communications
Finally, this section describes three visual observation experiments of cell-to-
cell communications to understand the diffusion and reaction characteristics.
First, pINV-112-R3 and pSND-1 were transformed into
E. coli
JM2.300 cells
to create sender cells. With IPTG induction, these sender cells emit cyan fluo-
rescence that serves as an easily identifiable marker due to the ECFP encoded
downstream of p(lac) on pINV-112-R3. In addition, the pSND-1 plasmid directs
α
1
In BL21-PRO cells, TetR (needed for controlled induction of the Tet promoter) exists on a plasmid, while in
DH5
α
TetR is part of the chromosomal DNA.
