pEYFP-C1 vector (V012016)

Price Information

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

Basic Vector Information

pEYFP-C1 vector 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.

References

• Uritu CM , Varganici CD , Ursu L , Coroaba A , Nicolescu A , Dascalu AI , Peptanariu D , Stan D , Constantinescu CA , Simion V , Calin M , Maier SS , Pinteala M , Barboiu M . Hybrid fullerene conjugates as vectors for DNA cell-delivery. J Mater Chem B. 2015 Mar 28;3(12):2433-2446. doi: 10.1039/c4tb02040e. Epub 2015 Feb 18. PMID: 32262120.
• Susa T, Kato T, Kato Y. Reproducible transfection in the presence of carrier DNA using FuGENE6 and Lipofectamine2000. Mol Biol Rep. 2008 Sep;35(3):313-9. doi: 10.1007/s11033-007-9088-0. Epub 2007 May 17. PMID: 17508267.
• Ben-Yair L, Slaaby R, Herman A, Cohen Y, Biener E, Moran N, Yoshimura A, Whittaker J, De Meyts P, Herman B, Gertler A. Preparation and expression of biologically active prolactin and growth hormone receptors and suppressor of cytokine signaling proteins 1, 2, 3, and 6 tagged with cyan and yellow fluorescent proteins. Protein Expr Purif. 2002 Aug;25(3):456-64. doi: 10.1016/s1046-5928(02)00044-x. PMID: 12182826.

pEYFP-C1 vector Sequence

LOCUS       Exported                4731 bp DNA     circular SYN 03-SEP-2024
DEFINITION  Vector for fusing EYFP to the N-terminus of a partner protein.
ACCESSION   .
VERSION     .
KEYWORDS    pEYFP-C1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4731)
  AUTHORS   Clontech
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 4731)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 4731)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..4731
                     /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"
     CDS             613..1329
                     /label=EYFP
                     /note="enhanced YFP"
     misc_feature    1330..1395
                     /label=MCS
                     /note="multiple cloning site"
     misc_feature    1404..1414
                     /label=stop codons
                     /note="stop codons"
                     /note="stop codons in all three reading frames"
     polyA_signal    1519..1640
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1647..2102)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2129..2233
                     /label=AmpR promoter
     promoter        2235..2592
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             2627..3418
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
     polyA_signal    3653..3700
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4029..4617
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"