Part:BBa_K3347009

Dehalogenase type 1, variant 2 + N His Tag Expression Block
Expression block for Dehalogenase type 1.variant 2.HisN (DeHa1.2.HisN gene; BBA_K3347008) for use in BL21-DE3 (pLysS) E. coli. This part's construction was inspired primarily from the standard pET-28 vector (T7 polymerase, RBS, and terminator) and adding DeHa1.2.HisN gene as a coding sequence. This sequence allows the expression of Dehalogenase type I with a amine-terminal 6xHis tag upon induction from IPTG using the T7 polymerase gene in the bacterial chromosome under expression control of LacUV5, an improved lac promoter.

Naming notation for Dehalogenase genes and expression blocks

Plasmid notation:
Represented by pXX_DeHa
Where XX can represent:
C: pSB1C3
A: pSB1A3
K: pSB1K3
KB: pSRKBB

Dehalogenase gene & Expression Block Notation
Represented by DeHaX.Y.Z
Where,
X is the gene type (1 or 2 ; referencing the two dehalogenase genes in D. acidovorans)
Y is the variant (native variant = 1, E. coli codon optimized = 2, etc.)
Z shows the tags/modifications on the protein (0 = no modifications, HisC = 6xHis tag on carboxyl terminal, HisN = 6xHis tag on amine terminal, gene, etc.)
Note: If Z is replaced by the 'gene' modifier, the Dehalogenase gene sequence is being referenced. In all other cases, the fully assembled expression block is referenced. Example: DeHa1.1.gene references BBa_K3347000 while DeHa1.1.0 references BBa_K3347001.

Example notations
pKB_DeHa2.2.HisN: Dehalogenase type II, variant 2 (E. coli optimized) with a N terminal 6xHis tag all inside the pSRKBB vector
pC_DeHa1.1.0: Dehalogenase type I, variant 1 with no further tags or modifications all inside the pSB1C3 vector

This expression system is used for large protein expression of DeHa1.2.HisN using the vector pSRKBB (pKB_DeHa1.2.HisN) inside the chassis BL21-DE3 (pLysS) E. coli
Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 220
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 388
  
• 1000
  COMPATIBLE WITH RFC[1000]