Lamp1-RFP vector (V007008)

Price Information

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V007008 Lamp1-RFP 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:
Lamp1-RFP
Antibiotic Resistance:
Kanamycin
Length:
5906 bp
Type:
Mammalian Expression
Replication origin:
ori
Selection Marker:
Neomycin (select with G418)
Copy Number:
High Copy
Promoter:
CMV
Cloning Method:
Restriction Enzyme
5' Primer:
CMV Forward
3' Primer:
DsRed1-N

Lamp1-RFP vector Map

Lamp1-RFP5906 bp60012001800240030003600420048005400SV40 promoterAmpR promoterf1 oriSV40 poly(A) signalmRFP1LAMP1CMV promoterCMV enhanceroriHSV TK poly(A) signalNeoR/KanR

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.

Lamp1-RFP vector Sequence

Copy Sequence

Download GenBank File(.gb)

LOCUS       40924_1594        5906 bp DNA     circular SYN 21-DEC-2021
DEFINITION  Mammalian expression of Lamp1 fused to RFP. Used for localization to
            the lysosome..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5906)
  AUTHORS   Sherer NM, Lehmann MJ, Jimenez-Soto LF, Ingmundson A, Horner SM, 
            Cicchetti G, Allen PG, Pypaert M, Cunningham JM, Mothes W
  TITLE     Visualization of retroviral replication in living cells reveals 
            budding into multivesicular bodies.
  JOURNAL   Traffic 2003 Nov;4(11):785-801.
  PUBMED    14617360
REFERENCE   2  (bases 1 to 5906)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 5906)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Traffic"; 
            date: "2003-11"; volume: "4(11)"; pages: "785-801"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5906
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(193..297)
                     /label=AmpR promoter
     rep_origin      324..779
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     polyA_signal    complement(786..907)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(1042..1716)
                     /codon_start=1
                     /label=mRFP1
                     /note="monomeric derivative of DsRed (Campbell et al.,
                     2002)"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     CDS             complement(1738..2958)
                     /codon_start=1
                     /label=LAMP1
                     /note="rat lysosome-associated membrane glycoprotein 1"
                     /translation="MAAPGARRPLLLLLLAGLAHSAPALFEVKDNNGTACIMASFSASF
                     LTTYEAGHVSKVSNMTLPASAEVLKNSSSCGEKNASEPTLAITFGEGYLLKLTFTKNTT
                     RYSVQHMYFTYNLSDTQFFPNASSKGPDTVDSTTDIKADINKTYRCVSDIRVYMKNVTI
                     VLWDATIQAYLPSSNFSKEETRCPQDQPSPTTGPPSPSPPLVPTNPSVSKYNVTGDNGT
                     CLLASMALQLNITYMKKDNTTVTRAFNINPSDKYSGTCGAQLVTLKVGNKSRVLELQFG
                     MNATSSLFFLQGVQLNMTLPDAIEPTFSTSNYSLKALQASVGNSYKCNSEEHIFVSKAL
                     ALNVFSVQVQAFRVESDRFGSVEECVQDGNNMLIPIAVGGALAGLVLIVLIAYLIGRKR
                     SHAGYQTI"
     promoter        complement(3033..3236)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     enhancer        complement(3237..3540)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(3715..4303)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     polyA_signal    complement(4632..4679)
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal (Cole and Stacy, 1985)"
     CDS             complement(4914..5705)
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     promoter        complement(join(5740..5906,1..191))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"