Difference between revisions of "Part:BBa K741012"

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The fusion protein crogfp is constantly expressed and the antisense RNA of cro is produced to inhibit the translation of the mRNA of crogfp after IPTG activated the promoter plac.  
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The fusion protein crogfp(λ phage Cro protein and GFP) is constantly expressed. The antisense RNA of cro is produced to inhibit the translation of the mRNA of crogfp after induced by IPTG.This part is used to measure the function of anticro.
  
 
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   width:767pt'>plac-RBS-anticro-T-pCon(0.856)-crogfp-T</td>
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   width:767pt'>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;plac-RBS-anticro-T-pCon(0.856)-crogfp-T</td>
 
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   <td colspan=10 height=19 class=xl24 style='height:14.25pt'>fluorescence
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   <td colspan=10 height=19 class=xl24 style='height:14.25pt'>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;fluorescence
 
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   <td colspan=10 height=19 class=xl24 style='height:14.25pt'>plac-RBS-anticro-T-pCon(0.856)-crogfp-T</td>
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   <td colspan=10 height=19 class=xl24 style='height:14.25pt'>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;plac-RBS-anticro-T-pCon(0.856)-crogfp-T</td>
 
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   <td colspan=10 height=19 class=xl24 style='height:14.25pt'>OD</td>
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   <td colspan=10 height=19 class=xl24 style='height:14.25pt'>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;OD</td>
 
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Latest revision as of 07:21, 3 October 2012

plac-anticro-T -pCon(0.856)-RBS-crogfp-T

The fusion protein crogfp(λ phage Cro protein and GFP) is constantly expressed. The antisense RNA of cro is produced to inhibit the translation of the mRNA of crogfp after induced by IPTG.This part is used to measure the function of anticro.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 124
    Illegal BglII site found at 1481
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 918

To test the function of antisense RNA to cro (anticro) we have constructed, we measure the fluorescence intensity of plac-anticroT-pcon(0.856)-crogfp and plac-cI-prm-anticroT-pcon(0.856)-crogfp.

USTC2012Result6 副本.png

At the beginning, the unit fluorescence intensity of the experimental groups with glucose added is the lowest, because glucose promotes the growth of E.coli., which makes the rate of fluorescence divided by OD lower than the other two groups.

About 45 minutes later, the unit fluorescence intensity of experimental groups with glucose added is higher than the blank group, because glucose represses the expression of plac, which reduces the transcription of anticro. Therefore, the translation of crogfp is repressed less and the unit fluorescence intensity is higher.

On the contrast, IPTG activates plac, which increases the transcription of anticro. Anticro binds to the mRNA crogfp and represses its translation.

Data

                                      plac-RBS-anticro-T-pCon(0.856)-crogfp-T
                                      fluorescence intensity
t/min Parallel 0 200mM glucose 400mM glucose 600mM glucose 800mM glucose 1000mM glucose 1000mM IPTG M9 medium
0 1 739.054 752.571 737.122 749.3 697.779 733.053 750.115 11.844
　 2 838.872 764.425 750.931 766.918 739.893 770.521 721.767 　
　 3 752.342 778.138 764.078 780.378 757.977 747.533 772.383 　
45 1 1121.065 1127.129 1132.739 1143.486 1104.684 1131.022 1070.284 12.897
　 2 1033.489 1079.895 1116.582 1157.266 1140.391 1144.527 1035.693 　
　 3 1040.658 1084.647 1158.246 1187.787 1190.048 1225.269 1027.305 　
90 1 1309.754 1503.889 1436.975 1510.864 1499.763 1546.764 1143.771 12.698
　 2 1290.674 1456.243 1444.75 1578.095 1411.988 1495.092 1101.698 　
　 3 1415.663 1317.712701 1542.16481 1490.563687 1490.115468 1510.510219 1064.565494 　
135 1 1294.433 1417.68 1502.504 1581.499 1520.589 1573.465 1263.79 12.945
　 2 1246.153 1466.956 1515.73 1548.78 1587.538 1559.81 1219.769 　
　 3 1305.599 1453.067 1460.94 1553.158 971.939 1603.972 1271.341 　
180 1 1438.159 1592.059 1595.049 1599.125 1573.181 1445.849 1206.695 11.793
　 2 1374.769 1432.318 1578.138 1643.468 1620.898 1525.023 1194.993 　
　 3 1311.343 1556.093 1551.538 1507.415 1458.318 1539.911 1135.83 　
                                           plac-RBS-anticro-T-pCon(0.856)-crogfp-T
                                                     OD
t/min Parallel 0 200mM glucose 400mM glucose 600mM glucose 800mM glucose 1000mM glucose 1000mM IPTG M9 medium
0 1 0.1052 0.1091 0.1117 0.1087 0.114 0.1052 0.1043 0.0374
　 2 0.108 0.1124 0.1179 0.1161 0.1107 0.1109 0.1036 　
　 3 0.1084 0.1991 0.1076 0.1124 0.112 0.1101 0.1006 　
45 1 0.1721 0.1657 0.1646 0.1601 0.1671 0.1805 0.1615 0.0412
　 2 0.161 0.1697 0.1717 0.1705 0.162 0.1633 0.1586 　
　 3 0.1593 0.1698 0.1633 0.1664 0.1676 0.1679 0.1518 　
90 1 0.2552 0.2604 0.2576 0.2532 0.2615 0.2659 0.2261 0.0379
　 2 0.2502 0.2569 0.2674 0.2752 0.2548 0.2672 0.2276 　
　 3 0.2545 0.2617 0.2587 0.2552 0.2694 0.2645 0.2176 　
135 1 0.2987 0.3379 0.3383 0.2969 0.3182 0.3331 0.2864 0.04
　 2 0.3163 0.3263 0.3316 0.3306 0.3245 0.3233 0.3154 　
　 3 0.3221 0.3347 0.3284 0.3196 0.3382 0.3358 0.3037 　
180 1 0.4115 0.412 0.4272 0.3793 0.4092 0.3986 0.3565 0.0406
　 2 0.4051 0.3951 0.411 0.421 0.3996 0.3746 0.374 　
　 3 0.4147 0.4107 0.3976 0.38 0.4029 0.3994 0.378