Difference between revisions of "Part:BBa K5043011"

Revision as of 12:38, 18 September 2024

amilGFP transcriptional unit

This part is a transcriptional unit, coding for amilGFP using Anderson promoter J23110, standard RBS B0034 and Terminator Luz7-T50. It was designed to verify protein production in Pseudomonas species.

Usage and Biology

Pseudomonas putida KT2440 [1] as well as Pseudomonas vancouverensis DSM8368 [2] show promising abilities as chassis for bioremediation of organic pollutants. [3, 4] In this context, we were interested in a low/medium constitutive expression system working in both bacteria. Hence, we decided to use Anderson promoter J23110, as it leads to medium expression in P. putida KT2440. [5] In addition, we chose RBS B0034 and Terminator Luz7-T50 both showing good efficiency in P. putida KT2440. [6, 7] As we could not find literature about these regulatory elements’ function in P. vancouverensis, amilGFP was used as reporter protein for qualitive assessment of protein production using this expression system. For this purpose, the shown composite part was cloned into pSEVA231-backbone. [8] Pseudomonas species were transformed using electroporatoration. Cell pellets of transformed liquid cultures exhibited no visible green colour under daylight, but green fluorescence under blue light.

As visible in the pictures, bacterial cultures bearing this composite part show green fluorescence, indicating GFP production. Thus, promoter J23110 and RBS B0034 both function in both P. putida KT2440 and P. vancouverensis DSM8368. The expression system is working.

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
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 161
Illegal AgeI site found at 622
1000
COMPATIBLE WITH RFC[1000]

@@ Line 3: / Line 3: @@
 <partinfo>BBa_K5043011 short</partinfo>
-Transcriptional unit coding for amilGFP using Anderson promoter J23110 and standard RBS B0034. This part was designed to verify protein production in Pseudomonas species.
+This part is a transcriptional unit, coding for amilGFP using Anderson promoter J23110, standard RBS B0034 and Terminator Luz7-T50. It was designed to verify protein production in Pseudomonas species.
-<!-- Add more about the biology of this part here
 ===Usage and Biology===
+Pseudomonas putida KT2440 [1] as well as Pseudomonas vancouverensis DSM8368 [2] show promising abilities as chassis for bioremediation of organic pollutants. [3, 4] In this context, we were interested in a low/medium constitutive expression system working in both bacteria. Hence, we decided to use Anderson promoter J23110, as it leads to medium expression in P. putida KT2440. [5] In addition, we chose RBS B0034 and Terminator Luz7-T50 both showing good efficiency in P. putida KT2440. [6, 7]
+As we could not find literature about these regulatory elements’ function in P. vancouverensis, amilGFP was used as reporter protein for qualitive assessment of protein production using this expression system. For this purpose, the shown composite part was cloned into pSEVA231-backbone. [8] Pseudomonas species were transformed using electroporatoration. Cell pellets of transformed liquid cultures exhibited no visible green colour under daylight, but green fluorescence under blue light.
+As visible in the pictures, bacterial cultures bearing this composite part show green fluorescence, indicating GFP production. Thus, promoter J23110 and RBS B0034 both function in both P. putida KT2440 and P. vancouverensis DSM8368. The expression system is working.
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