Difference between revisions of "Part:BBa K3351000"

(Characterization by NWU-CHINA-A 2021)
 
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===Characterization by NWU-CHINA-A 2021===
 
===Characterization by NWU-CHINA-A 2021===
 
<p>We used this part in our composite part BBa_K3722007, BBa_K3722008 and BBa_K3722009 to express tryptophanase (TnaA, BBa_K3722000), monooxygenase (MaFMO, BBa_K3722002) and tryptophan (Trp) halogenase (SttH, BBa_K3722003). All of them were constructed into pET-28a and transformed to E.coli (DE3) for expression.</p>
 
<p>We used this part in our composite part BBa_K3722007, BBa_K3722008 and BBa_K3722009 to express tryptophanase (TnaA, BBa_K3722000), monooxygenase (MaFMO, BBa_K3722002) and tryptophan (Trp) halogenase (SttH, BBa_K3722003). All of them were constructed into pET-28a and transformed to E.coli (DE3) for expression.</p>
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[[File:T--NWU-CHINA-A--SDS-PAGE result of TnaA, SttH, MaFMO.jpg|350px|center|]]
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[[File:T--NWU-CHINA-A--SDS-PAGE result of TnaA, SttH, MaFMO.jpg|500px|center|]]
 
<p><center><b>Figure 1. SDS-PAGE result of TnaA, SttH, MaFMO. Lane on the far left is protein weight marker. Lane 1, tryptophanase (TnaA). Lane 2, tryptophan halogenase (SttH). Lane 3, monooxygenase (MaFMO).</b></center></p>
 
<p><center><b>Figure 1. SDS-PAGE result of TnaA, SttH, MaFMO. Lane on the far left is protein weight marker. Lane 1, tryptophanase (TnaA). Lane 2, tryptophan halogenase (SttH). Lane 3, monooxygenase (MaFMO).</b></center></p>
 
[[File:T--NWU-CHINA-A--SDS-PAGE analysis of MaFMO.jpg|500px|center|]]
 
[[File:T--NWU-CHINA-A--SDS-PAGE analysis of MaFMO.jpg|500px|center|]]
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<p><center><b>1: negative control  2: 9h  3:10.5h  4:12h  5:13.5h  6:15h  7:16.5h  8: 18h  9: 19.5h
 
<p><center><b>1: negative control  2: 9h  3:10.5h  4:12h  5:13.5h  6:15h  7:16.5h  8: 18h  9: 19.5h
 
</b></center></p>
 
</b></center></p>
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===Characterization by NWU-CHINA-A 2022===
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<p>We used this part in our composite part BBa_K4199005, BBa_K4199006 and BBa_K4199007 to express ChuA(BBa_K4199001), HrtR(BBa_K4199002). All of them were constructed into pET-28a and transformed to E.coli BL21(DE3) for expression.</p>
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[[File:NWU-CHINA-A--SDS-PAGE result of supernatant.jpg|500px|center|]]
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<p><center><b>Figure 1. SDS-PAGE result of supernatant of bacteria broken by ultrasound.</b></center></p>
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[[File:NWU-CHINA-A--SDS-PAGE result of precipitate.jpg|500px|center|]]
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<p><center><b>Figure 2. SDS-PAGE results of resuspended precipitate after ultrasonic fragmentation of bacteria.</b></center></p>
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<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Latest revision as of 08:49, 11 October 2022


A T7 promoter from pet28a plasmid.

Summary

It is a strong promoter from T7 bacteriophage that can specifically react to T7RNA polymerase. It is a sequence that initiates the transcription of T7 bacteriophage gene.

Characterization by NWU-CHINA-A 2021

We used this part in our composite part BBa_K3722007, BBa_K3722008 and BBa_K3722009 to express tryptophanase (TnaA, BBa_K3722000), monooxygenase (MaFMO, BBa_K3722002) and tryptophan (Trp) halogenase (SttH, BBa_K3722003). All of them were constructed into pET-28a and transformed to E.coli (DE3) for expression.

T--NWU-CHINA-A--SDS-PAGE result of TnaA, SttH, MaFMO.jpg

Figure 1. SDS-PAGE result of TnaA, SttH, MaFMO. Lane on the far left is protein weight marker. Lane 1, tryptophanase (TnaA). Lane 2, tryptophan halogenase (SttH). Lane 3, monooxygenase (MaFMO).

T--NWU-CHINA-A--SDS-PAGE analysis of MaFMO.jpg

Figure 2. SDS-PAGE analysis of MaFMO. Induction condition: LB media, OD600 0.6 ~ 0.8, IPTG 0.1 mM, 18°C and 200 rpm.

0:negative control 1:4h 2:5h 3:6h 4:7h 5:8h 6:9h 7:10h 8:11h

T--NWU-CHINA-A--SDS-PAGE analysis of TnaA.jpg

Figure 3. SDS-PAGE analysis of TnaA. Induction condition: LB media, OD600 0.6 ~ 0.8, IPTG 0.1 mM, 37 °C and 200 rpm.

1: negative control 2: 9h 3:10.5h 4:12h 5:13.5h 6:15h 7:16.5h 8: 18h 9: 19.5h


Characterization by NWU-CHINA-A 2022

We used this part in our composite part BBa_K4199005, BBa_K4199006 and BBa_K4199007 to express ChuA(BBa_K4199001), HrtR(BBa_K4199002). All of them were constructed into pET-28a and transformed to E.coli BL21(DE3) for expression.

NWU-CHINA-A--SDS-PAGE result of supernatant.jpg

Figure 1. SDS-PAGE result of supernatant of bacteria broken by ultrasound.

NWU-CHINA-A--SDS-PAGE result of precipitate.jpg

Figure 2. SDS-PAGE results of resuspended precipitate after ultrasonic fragmentation of bacteria.



Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]