Basic Vector Information
- Vector Name:
- pMKO.1 puro
- Antibiotic Resistance:
- Ampicillin
- Length:
- 6340 bp
- Type:
- Mammalian Expression, Retroviral, RNAi
- Replication origin:
- ori
- Selection Marker:
- Puromycin
- Copy Number:
- High Copy
- Promoter:
- U6
- Cloning Method:
- Restriction Enzyme
- 5' Primer:
- pLXSN 5'
- 3' Primer:
- pBABE-3
pMKO.1 puro vector Vector Map
Plasmid Resuspension 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 ℃.
pMKO.1 puro vector Sequence
LOCUS 40924_31055 6340 bp DNA circular SYN 13-MAY-2021
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6340)
AUTHORS Stewart SA, Dykxhoorn DM, Palliser D, Mizuno H, Yu EY, An DS,
Sabatini DM, Chen IS, Hahn WC, Sharp PA, Weinberg RA, Novina CD
TITLE Lentivirus-delivered stable gene silencing by RNAi in primary cells.
JOURNAL RNA 2003 Apr;9(4):493-501.
PUBMED 12649500
REFERENCE 2 (bases 1 to 6340)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 6340)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "RNA"; date:
"2003-04"; volume: "9(4)"; pages: "493-501"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..6340
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 1..241
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
rep_origin 490..625
/label=SV40 ori
/note="SV40 origin of replication"
CDS 649..1245
/codon_start=1
/label=PuroR
/note="puromycin N-acetyltransferase"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
LTR 1444..1869
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
Moloney murine leukemia virus"
primer_bind complement(1972..1994)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
promoter 2135..2464
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
primer_bind 2486..2505
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind complement(2586..2603)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2757..3345)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(3519..4376)
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(4377..4481)
/label=AmpR promoter
primer_bind 4549..4567
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
enhancer 4667..5046
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 5047..5250
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
LTR 5251..5426
/label=5' LTR (truncated)
/note="truncated long terminal repeat from Moloney murine
sarcoma virus"
misc_feature 5489..5688
/label=MMLV Psi
/note="packaging signal of Moloney murine leukemia virus
(MMLV)"
CDS 5889..6305
/codon_start=1
/label=gag (truncated)
/note="truncated Moloney murine leukemia virus (MMLV) gag
gene lacking the start codon"
/translation="GQTVTTPLSLTLGHWKDVERIAHNQSVDVKKRRWVTFCSAEWPTF
NVGWPRDGTFNRDLITQVKIKVFSPGPHGHPDQVPYIVTWEALAFDPPPWVKPFVHPKP
PPPLPPSAPSLPLEPPRSTPPRSSLYPALTPSLGA"
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