Part:BBa_K3351000

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.

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).

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

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.

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

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]