Basic Vector Information
pNZ8148 contains an origin of replication (ORI), the gene for the resistance to chloramphenicol, two genes for the replication proteins repA and repC, the nisin-inducible promoter (P nisA), and the transcription terminator (T). The gene, tagged with a Strep-tag II (STREP) followed by a stop codon (*), can be inserted between the necessary NcoI site and another endonuclease site from the multicloning site (MCS) such as PstI, SphI, KpnI, SpeI, XbaI, SacI and HindIII. The replicons of the vectors pNZ8008, pNZ8148, pNZ8149 and pNZ8150 are identical and came originally from the Lactococcus lactis plasmid pSH71. However, this replicon has a broad host range. Plasmids with this replicon can replicate in many Gram-positive bacteria, such as Lactobacillus plantarum and Streptococcus thermophilus. pNZ8148 – In this vector the nisA promoter is followed by an NcoI site for translational fusions at the ATG. It contains a terminator after the MCS. Sequence adaptation for cloning in NcoI can result in a change in the second amino acid of a protein (Mierau and Kleerebezem, 2005).
| Basic Vector Information | |||
|---|---|---|---|
| Vector Name | pNZ8148 | Antibiotic Resistance | Chloramphenicol |
| Length | 3167 bp | Type | expression plasmid of Lactobacilli |
| Promoter | PnisA | ||
pNZ8148 vector Vector Map
Plasmid Resuspension protocol:
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5.Store the plasmid at -20 ℃.
pNZ8148 vector Sequence
LOCUS Exported 3167 bp ds-DNA circular SYN 17-JUN-2022
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS pNZ8148
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 3167)
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..3167
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 5..188
/note="Pnis"
/note="Nisin A promoter region"
terminator 393..445
/note="T"
CDS 788..997
/codon_start=1
/note="repC"
/note="Replication gene C"
/translation="MGGKEANFASVLRPPIKCRVPIFVPKTLYPNWLKGLRGFSIANES
PTFSPTFFINLYLSSFIFVFMITK"
CDS 1266..1964
/codon_start=1
/note="repA"
/note="Replication gene A"
/translation="MAIKNTKARNFGFLLYPDSIPNDWKEKLESLGVSMAVSPLHDMDE
KKDKDTWNSSDVIRNGKHYKKPHYHVIYIARNPVTIESVRNKIKRKLGNSSVAHVEILD
YIKGSYEYLTHESKDAIAKNKHIYDKKDILNINDFDIDRYITLDESQKRELKNLLLDIV
DDYNLVNTKDLMAFIRLRGAEFGILNTNDVKDIVSTNSSAFRLWFEGNYQCGYRASYAK
VLDAETGEIK"
CDS 2424..3074
/codon_start=1
/note="cm"
/note="chloramphenicol resistant gene"
/translation="MNFNKIDLDNWKRKEIFNHYLNQQTTFSITTEIDISVLYRNIKQE
GYKFYPAFIFLVTRVINSNTAFRTGYNSDGELGYWDKLEPLYTIFDGVSKTFSGIWTPV
KNDFKEFYDLYLSDVEKYNGSGKLFPKTPIPENAFSLSIIPWTSFTGFNLNINNNSNYL
LPIITAGKFINKGNSIYLPLSLQVHHSVCDGYHAGLFMNSIQELSDRPNDWLL"
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