Cart 2

pDawn-mCherry-Amp vector (V014895)

Price Information

Cat No. Plasmid Name Availability Add to cart
V014895 pDawn-mCherry-Amp In stock, 1 week for quality controls

Buy one, get one free!

Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.

Basic Vector Information

Vector Name:
pDawn-mCherry-Amp
Antibiotic Resistance:
Ampicillin
Length:
8089 bp
Replication origin:
ori

pDawn-mCherry-Amp vector Map

Copy Sequence View Map
pDawn-mCherry-Amp8089 bp400800120016002000240028003200360040004400480052005600600064006800720076008000f1 oriAmpRAmpR promoteroribomropTranscriptional regulatory protein FixJlacIq promoterlambda repressorssrA tag (LVA)rrnB T1 terminatorT7Te terminatorRBS6xHisthrombin sitemCherry6xHisT7 terminator

Plasmid 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 ℃.

6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it

General Plasmid Transform Protocol

1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.

2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.

3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.

4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.

5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.

pDawn-mCherry-Amp vector Sequence

LOCUS       V014895                 8089 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  Exported.
ACCESSION   V014895
VERSION     V014895
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 8089)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8089
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      12..467
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     CDS             complement(591..1448)
                     /label="AmpR"
                     /note="beta-lactamase"
     promoter        complement(1449..1553)
                     /label="AmpR promoter"
     rep_origin      1679..2267
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     misc_feature    complement(2453..2595)
                     /label="bom"
                     /note="basis of mobility region from pBR322"
     CDS             complement(2700..2888)
                     /label="rop"
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
     CDS             complement(3692..4306)
                     /gene="fixJ"
                     /label="Transcriptional regulatory protein FixJ"
                     /note="Transcriptional regulatory protein FixJ from
                     Bradyrhizobium diazoefficiens (strain JCM 10833 / BCRC
                     13528 / IAM 13628 / NBRC 14792 / USDA 110). Accession#:
                     P23221"
     promoter        complement(5446..5523)
                     /label="lacIq promoter"
                     /note="In the lacIq allele, a single base change in the
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             6089..6799
                     /label="lambda repressor"
                     /note="phage lambda repressor"
     CDS             6800..6832
                     /label="ssrA tag (LVA)"
                     /note="C-terminal peptide that mediates degradation in
                     bacteria through the ClpXP and ClpAP proteases (McGinness
                     et al., 2006)"
     terminator      6880..6951
                     /label="rrnB T1 terminator"
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      6967..6994
                     /label="T7Te terminator"
                     /note="phage T7 early transcription terminator"
     RBS             7070..7092
                     /label="RBS"
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             7111..7128
                     /label="6xHis"
                     /note="6xHis affinity tag"
     CDS             7138..7155
                     /label="thrombin site"
                     /note="thrombin recognition and cleavage site"
     CDS             7201..7908
                     /label="mCherry"
                     /note="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
     CDS             7933..7950
                     /label="6xHis"
                     /note="6xHis affinity tag"
     terminator      8017..8064
                     /label="T7 terminator"
                     /note="transcription terminator for bacteriophage T7 RNA
                     polymerase"