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pKaede-S1 vector (V011992)

Price Information

Cat No. Plasmid Name Availability Add to cart
V011992 pKaede-S1 In stock, 1 week for quality controls

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

Basic Vector Information

Vector Name:
pKaede-S1
Antibiotic Resistance:
Ampicillin
Length:
3361 bp
Type:
Fluorescent Protein Genes & Plasmids
Replication origin:
ori
Source/Author:
MBL International
Copy Number:
High copy number
5' Primer:
M13 fwd
3' Primer:
M13 rev

pKaede-S1 vector Map

Copy Sequence View Map
pKaede-S13361 bp6001200180024003000AmpR promoterAmpRoriCAP binding sitelac promoterlac operatorM13 revKaedeM13 fwd

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.

pKaede-S1 vector Sequence

LOCUS       40924_26521        3361 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Vector for expressing Kaede in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3361)
  AUTHORS   MBL International
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 3361)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..3361
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        96..200
                     /label=AmpR promoter
     CDS             201..1058
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      1232..1820
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    2108..2129
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        2144..2174
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2182..2198
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             2265..2942
                     /codon_start=1
                     /label=Kaede
                     /note="Kaede green-to-red photoconvertible fluorescent
                     protein (Ando et al., 2002)"
                     /translation="MVSLIKPEMKIKLLMEGNVNGHQFVIEGDGKGHPFEGKQSMDLVV
                     KEGAPLPFAYDILTTAFHYGNRVFAKYPDHIPDYFKQSFPKGFSWERSLMFEDGGVCIA
                     TNDITLKGDTFFNKVRFDGVNFPPNGPVMQKKTLKWEASTEKMYLRDGVLTGDITMALL
                     LKGDVHYRCDFRTTYKSRQEGVKLPGYHFVDHCISILRHDKDYNEVKLYEHAVAHSGLP
                     DNVK"
     primer_bind     complement(2967..2983)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"