pKatushka2S-N vector (V011989)

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Price Information

Cat No.	Plasmid Name	Availability	Add to cart
V011989	pKatushka2S-N	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:: pKatushka2S-N

Antibiotic Resistance:: Kanamycin

Length:: 4720 bp

Type:: Fluorescent Protein Genes & Plasmids

Replication origin:: ori

Source/Author:: Luker KE, Pata P, Shemiakina II, Pereverzeva A, Stacer AC, Shcherbo

Selection Marker:: Neomycin/G418(Geneticin)

Copy Number:: High copy number

pKatushka2S-N vector Map

Copy Sequence View Map

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.

pKatushka2S-N vector Sequence

LOCUS       40924_26566        4720 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Vector for fusing Katushka2S to the C-terminus of a partner protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4720)
  AUTHORS   Luker KE, Pata P, Shemiakina II, Pereverzeva A, Stacer AC, Shcherbo 
            DS, Pletnev VZ, Skolnaja M, Lukyanov KA, Luker GD, Pata I, Chudakov 
            DM.
  TITLE     Comparative study reveals better far-red fluorescent protein for 
            whole body imaging.
  JOURNAL   Sci Rep 2015;5:10332.
  PUBMED    26035795
REFERENCE   2  (bases 1 to 4720)
  AUTHORS   Evrogen
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 4720)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Rep"; 
            date: "2015"; volume: "5"; pages: "10332"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..4720
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     misc_feature    591..671
                     /label=MCS
                     /note="multiple cloning site"
     CDS             679..1383
                     /codon_start=1
                     /label=Katushka2S
                     /note="improved version of the oligomeric far-red
                     fluorescent protein TurboFP635/Katushka (Luker et al., 
                     2015)"
                     /translation="MVGEDSVLITENMHMKLYMEGTVNDHHFKCTSEGEGKPYEGTQTM
                     KIKVVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERITTYEDGG
                     VLTATQDTSLQNGCLIYNVKINGVNFPSNGPVMQKKTLGWEASTEMLYPADSGLRGHAQ
                     MALKLVGGGYLHCSLKTTYRSKKPAKNLKMPGFYFVDRRLERIKEADKETYVEQHEMAV
                     ARYCDLPSKLGHS"
     polyA_signal    1508..1629
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1636..2091)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2118..2222
                     /label=AmpR promoter
     promoter        2224..2581
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             2616..3407
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    3642..3689
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4018..4606
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"