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pDONR207 SARS-CoV-2 NSP3 vector (V006598)

Price Information

Cat No. Plasmid Name Availability Add to cart
V006598 pDONR207 SARS-CoV-2 NSP3 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:
pDONR207 SARS-CoV-2 NSP3
Antibiotic Resistance:
Gentamicin
Length:
9211 bp
Type:
Gateway-compatible Entry vector
Replication origin:
ori
Promoter:
Pc
Cloning Method:
Gateway Cloning
5' Primer:
NA

pDONR207 SARS-CoV-2 NSP3 vector Map

Copy Sequence View Map
pDONR207 SARS-CoV-2 NSP39211 bp400800120016002000240028003200360040004400480052005600600064006800720076008000840088009200rrnB T2 terminatorrrnB T1 terminatorattL5attL2pENTR-RKan-RGmRPc promoterori

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.

pDONR207 SARS-CoV-2 NSP3 vector Sequence

LOCUS       40924_15090        9211 bp DNA     circular SYN 13-MAY-2021
DEFINITION  Gateway-compatible Entry vector, with insert of NSP3 gene's CDS from
            SARS-CoV-2 isolate Wuhan-Hu-1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9211)
  AUTHORS   Kim DK, Knapp JJ, Kuang D, Chawla A, Cassonnet P, Lee H, 
            Sheykhkarimli D, Samavarchi-Tehrani P, Abdouni H, Rayhan A, Li R, 
            Pogoutse O, Coyaud E, van der Werf S, Demeret C, Gingras AC, Taipale
            M, Raught B, Jacob Y, Roth FP
  TITLE     A Comprehensive, Flexible Collection of SARS-CoV-2 Coding Regions.
  JOURNAL   G3 (Bethesda). 2020 Aug 6. pii: g3.120.401554. doi: 
            10.1534/g3.120.401554.
  PUBMED    32763951
REFERENCE   2  (bases 1 to 9211)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 9211)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "G3 
            (Bethesda). 2020 Aug 6. pii: g3.120.401554. doi: 
            10.1534/g3.120.401554."
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..9211
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(56..83)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(175..261)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     protein_bind    315..410
                     /label=attL5
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    complement(6255..6354)
                     /label=attL2
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(6466..6485)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(6544..6563)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(7140..7670)
                     /codon_start=1
                     /label=GmR
                     /note="gentamycin acetyltransferase"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(7859..7887)
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     rep_origin      8544..9132
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"