Part:BBa_K2273104

AmyE_RFC25 version of B. subtilis alpha-amylase without its native signal peptide

AmyE_RFC25 is used in the 2017 TU Dresden iGEM project [http://2017.igem.org/Team:TU_Dresden EncaBcillus - It's a trap!].

The AmyE_EV (BBa_K2273103) which was amplified from AmyE_RFC25 is used as gene of interest for proof of concept of the Signal Peptide Toolbox of [http://2017.igem.org/Team:TU_Dresden iGEM Team TU Dresden 2017 (EncaBcillus - It's a trap!)].

The AmyE_RFC25 version of B. subtilis alpha-amylase (Uniprot [http://www.uniprot.org/uniprot/P00691 P00691]) without its first 33 amino acids (Brockmeier et al, 2006) which code for the native signal peptide would naturally be secreted into the supernatant to degrade extracellular starch. However, as its native signal peptide has been cut off, it lacks the naturally occuring secretion capability.

This part has the following prefix and suffix which includes a strong Shine Dalgarno Sequence (SD):

Prefix with	EcoRI, NotI, XbaI and SD	5'-GAATTCGCGGCCGCTTCTAGAGAAGGAGTGTCAAGA-3'
Suffix with	SpeI, NotI and PstI	5'-TACTAGTAGCGGCCGCTGCAG-3'

Sites of restriction enzymes generating compatible overhangs are indicated by sharing one color. (EcoRI and PstI are marked in blue, NotI in green, XbaI and SpeI are red. Additionally, the Shine Dalgarno Sequence is marked in silver.)

Sequence and Features

The DNA sequence of this part has been verified via sequencing before it was sent in.

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]