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pQE-100 DoubleTag vector (V010809)

Price Information

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V010809 pQE-100 DoubleTag In stock, instant shipping

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

Basic Vector Information

Vector Name:
pQE-100 DoubleTag
Antibiotic Resistance:
Ampicillin
Length:
3512 bp
Type:
Qiagen Vectors
Replication origin:
ori
Source/Author:
Qiagen
Copy Number:
High copy number
Growth Strain(s):
DH10B
Growth Temperature:
37℃

pQE-100 DoubleTag vector Map

Copy Sequence View Map
pQE-100 DoubleTag3512 bp6001200180024003000T5 promoterlac operatorRBSATG6xHisMCSTag-100stop codonslambda t0 terminatorCmRrrnB T1 terminatorbomoriAmpRAmpR promoter

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.

pQE-100 DoubleTag vector Sequence

LOCUS       pQE-100_DoubleTa        3512 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Bacterial vector for expressing proteins with an N-terminal 6xHis 
            tag and a C-terminal Tag-100 tag.
ACCESSION   .
VERSION     .
KEYWORDS    pQE-100 DoubleTag
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3512)
  AUTHORS   Qiagen
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 3512)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..3512
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        10..54
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA
                     polymerase, with embedded lac operator"
     protein_bind    62..78
                     /label=lac repressor encoded by lacI binding site
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="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)."
     RBS             101..106
                     /note="ribosome binding site"
     CDS             115..117
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             127..144
                     /label=6xHis
                     /note="6xHis affinity tag"
     misc_feature    145..184
                     /label=MCS
                     /note="MCS"
                     /note="multiple cloning site"
     CDS             193..228
                     /label=Tag-100
                     /note="epitope tag derived from MAP kinase 2"
     misc_feature    243..253
                     /label=stop codons
                     /note="stop codons"
                     /note="stop codons in all three reading frames"
     terminator      259..353
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             397..1053
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     terminator      1121..1207
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     misc_feature    1362..1502
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(1688..2276)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2450..3307)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(3308..3412)
                     /label=AmpR promoter