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phRG(R2.2) vector (V005502) Gene synthesis in phRG(R2.2) backbone

Price Information

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V005502 phRG(R2.2) In stock, instant shipping

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

Basic Vector Information

Vector Name:
phRG(R2.2)
Antibiotic Resistance:
Ampicillin
Length:
4304 bp
Type:
Reporter vector
Replication origin:
ori
Source/Author:
Almond BD, Chauvin F, Kenefick KB.
Growth Strain(s):
Top10

phRG(R2.2) vector Map

Copy Sequence View Map
phRG(R2.2)4304 bp600120018002400300036004200multiple cloning regionhRluchCL1hPESTSV40 poly(A) signalreporter vector primer 4 (RVPrimer4)ColE1-derivedoriAmpRAmpR promoterf1 oripoly(A) signalpause site

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.

phRG(R2.2) vector Sequence

LOCUS       40924_24947        4304 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Reporter vector phRG(R2.2), complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4304)
  AUTHORS   Almond BD, Chauvin F, Kenefick KB.
  TITLE     Rapid Response (TM) Reporter Vectors Technical Manual, TM242
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4304)
  AUTHORS   Almond BD, Chauvin F, Kenefick KB.
  TITLE     Direct Submission
  JOURNAL   Submitted (26-NOV-2003) Scientific Communications, Promega 
            Corporation, 2800 Woods Hollow Rd., Madison, WI 53711, USA
REFERENCE   3  (bases 1 to 4304)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 4304)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (26-NOV-2003) Scientific Communications, Promega Corporation, 2800 
            Woods Hollow Rd., Madison, WI 53711, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..4304
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..58
                     /label=multiple cloning region
                     /note="multiple cloning region"
     CDS             88..1020
                     /codon_start=1
                     /label=hRluc
                     /note="Renilla luciferase"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     CDS             1027..1074
                     /codon_start=1
                     /label=hCL1
                     /note="non-ORF yeast peptide conferring ubiquitin-dependent
                     degradation"
                     /translation="ACKNWFSSLSHFVIHL"
     CDS             1078..1197
                     /codon_start=1
                     /label=hPEST
                     /note="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     polyA_signal    complement(1267..1388)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     1547..1566
                     /label=reporter vector primer 4 (RVPrimer4)
                     /note="reporter vector primer 4 (RVPrimer4)"
     rep_origin      1804
                     /label=ColE1-derived
                     /note="ColE1-derived"
     rep_origin      complement(1807..2395)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2569..3426)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(3427..3531)
                     /label=AmpR promoter
     rep_origin      3558..4013
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     polyA_signal    4144..4192
                     /label=poly(A) signal
                     /note="synthetic polyadenylation signal"
     misc_feature    4206..4297
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from
                     the human alpha-2 globin gene"