Difference between revisions of "Part:BBa K2203004"
Melekcabani (Talk | contribs) |
Melekcabani (Talk | contribs) |
||
| Line 5: | Line 5: | ||
This composite part consists of a T7 promoter (BBa_J64997) in front of the toehold, a lacZalpha (BBa_I732006) downstream of it and a T7 terminator (BBa_K731721) at the end of it. | This composite part consists of a T7 promoter (BBa_J64997) in front of the toehold, a lacZalpha (BBa_I732006) downstream of it and a T7 terminator (BBa_K731721) at the end of it. | ||
Sequence of toehold 27B detecting a part of the Zika virus. This part can now be used in combination with cell-free extracts to detect the presence of Zika virus in a sample. | Sequence of toehold 27B detecting a part of the Zika virus. This part can now be used in combination with cell-free extracts to detect the presence of Zika virus in a sample. | ||
| + | |||
The reporter system in our toeholds(K2203004, K2203005 for zika detection and K2203002, K2203006 for hepatitis C detection works as follows: | The reporter system in our toeholds(K2203004, K2203005 for zika detection and K2203002, K2203006 for hepatitis C detection works as follows: | ||
| + | |||
- M15 cells have a lacZ delta mutation which makes them encode a form of beta-galactosidase lacking residues 11-41. | - M15 cells have a lacZ delta mutation which makes them encode a form of beta-galactosidase lacking residues 11-41. | ||
| + | |||
- Beta-galactosidase produced without those residues is missing a small part and is thus not functional. | - Beta-galactosidase produced without those residues is missing a small part and is thus not functional. | ||
| + | |||
- Upon the annealing of a trigger zika sequence to the toehold switch, the toehold unfolds and the lacZalpha coding sequence is translated. | - Upon the annealing of a trigger zika sequence to the toehold switch, the toehold unfolds and the lacZalpha coding sequence is translated. | ||
| + | |||
- The production of the lacZaplpha part and the indigenous presence of the lacZ omega part in M15 cells allows for alpha complementation and production of a functional beta-galactosidase that can be detected by the addition of a colorimetric substrate CPRG Chlorophenol red-β-D-galactopyranoside. | - The production of the lacZaplpha part and the indigenous presence of the lacZ omega part in M15 cells allows for alpha complementation and production of a functional beta-galactosidase that can be detected by the addition of a colorimetric substrate CPRG Chlorophenol red-β-D-galactopyranoside. | ||
===Characterization of the 27B_Zika Toehold in T7M15 lysate in the presence of a single stranded DNA trigger=== | ===Characterization of the 27B_Zika Toehold in T7M15 lysate in the presence of a single stranded DNA trigger=== | ||
<html> | <html> | ||
| + | <center> | ||
| + | |||
<figure> | <figure> | ||
<img src="https://static.igem.org/mediawiki/parts/9/9b/27B.png" height="480" width="640"> | <img src="https://static.igem.org/mediawiki/parts/9/9b/27B.png" height="480" width="640"> | ||
<figcaption> <center>Repeats of T7LacZalpha in T7M15 lysate </center></figcaption> | <figcaption> <center>Repeats of T7LacZalpha in T7M15 lysate </center></figcaption> | ||
</figure> | </figure> | ||
| + | </center> | ||
| + | |||
</html> | </html> | ||
Revision as of 11:43, 29 October 2017
ZIKV_27B_lacZalpha
This composite part consists of a T7 promoter (BBa_J64997) in front of the toehold, a lacZalpha (BBa_I732006) downstream of it and a T7 terminator (BBa_K731721) at the end of it. Sequence of toehold 27B detecting a part of the Zika virus. This part can now be used in combination with cell-free extracts to detect the presence of Zika virus in a sample.
The reporter system in our toeholds(K2203004, K2203005 for zika detection and K2203002, K2203006 for hepatitis C detection works as follows:
- M15 cells have a lacZ delta mutation which makes them encode a form of beta-galactosidase lacking residues 11-41.
- Beta-galactosidase produced without those residues is missing a small part and is thus not functional.
- Upon the annealing of a trigger zika sequence to the toehold switch, the toehold unfolds and the lacZalpha coding sequence is translated.
- The production of the lacZaplpha part and the indigenous presence of the lacZ omega part in M15 cells allows for alpha complementation and production of a functional beta-galactosidase that can be detected by the addition of a colorimetric substrate CPRG Chlorophenol red-β-D-galactopyranoside.
Characterization of the 27B_Zika Toehold in T7M15 lysate in the presence of a single stranded DNA trigger
Incubation reaction of 27B_Zika Toehold in T7M15 lysate in the presence of a single stranded DNA trigger
Right: Toehold LacZ alpha with no trigger added: the alpha part cannot be translated
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]