pMD18-T Simple vector (V017515)

Price Information

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V017515 pMD18-T Simple 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

pMD18-T Simple Vector is a specialized vector for efficient cloning of PCR products (TA Cloning). This vector is converted from pUC18 vector by inserting an EcoRV recognition site between XbaI and Sa1I recognition sites at the multiple cloning site of pUC18 vector, and then adding a “T” at the 3' end of both sides after enzymatic digestion with EcoR V. The pMD*18-T Vector is a special vector for highly efficient cloning of PCR products (TA Cloning). Because most of the thermophilic DNA polymerase PCR reaction has the characteristic of adding an “A” at the 3' end of the PCR product, the use of this product can greatly improve the efficiency of ligation and cloning of PCR products. Since this vector is constructed on the basis of pUC18 vector, it has the same function as pUC18 vec

Vector Name:
pMD18-T Simple
Antibiotic Resistance:
Ampicillin
Length:
2692 bp
Type:
TA Cloning Vector
Source/Author:
Long JY, Chen YH, Xia JR.
Copy Number:
High Copy
Promoter:
lac
5' Primer:
M13 fwd
3' Primer:
M13 rev
Growth Strain(s):
suitable for use in most common commercial host cells like TOP10, DH5α, TOP10F and JM109

pMD18-T Simple vector Map

pMD18-T Simple2692 bp600120018002400lacZ-alphalac operatorlac promoterCAP binding siteoriAmpRAmpR 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.

References

  • Long JY, Chen YH, Xia JR. First Report of a Group 16SrI Phytoplasma Associated with Amaranthus hypochondriacus Cladodes in China. Plant Dis. 2011 Jul;95(7):871. doi: 10.1094/PDIS-03-11-0219. PMID: 30731728.

pMD18-T Simple vector Sequence

LOCUS       Exported                2692 bp DNA     circular SYN 20-JAN-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2692)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..2692
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(146..475)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITNSSSVPGDPLEISSTCRHASLALAVVLQRRDWENPGVTQL
                     NRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTI
                     CSDAA"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(471..487)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    495..511
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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)."
     promoter        complement(519..549)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    564..585
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(873..1461)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1632..2492)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(2493..2597)
                     /gene="bla"
                     /label=AmpR promoter