LOCUS       sequence-203919-        7236 bp    DNA     circular SYN 15-APR-2025
DEFINITION  creating stable line with GFP and G418 resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7236)
  AUTHORS   Liao J, Wei Q, Fan J, Zou Y, Song D, Liu J, Liu F, Ma C, Hu X, Li L,
            Yu Y, Qu X, Chen L, Yu X, Zhang Z, Zhao C, Zeng Z, Zhang R, Yan S, 
            Wu T, Wu X, Shu Y, Lei J, Li Y, Zhang W, Wang J, Reid RR, Lee MJ, 
            Huang W, Wolf JM, He TC, Wang J
  TITLE     Characterization of retroviral infectivity and superinfection 
            resistance during retrovirus-mediated transduction of mammalian 
            cells.
  JOURNAL   Gene Ther. 2017 Jun;24(6):333-341. doi: 10.1038/gt.2017.24. Epub 
            2017 May 4.
  PUBMED    28387759
REFERENCE   2  (bases 1 to 7236)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/gt.2017.24"; journalName: "Gene Ther"; date: "2017-06"; 
            volume: "24"; issue: "6"; pages: "333-341"
COMMENT     Sequence Label: pRV-GN
FEATURES             Location/Qualifiers
     source          1..7236
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             192..708
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    772..1113
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             1180..1596
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1526..1548
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1564..1580
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             1607..2407
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MAMIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQG
                     RPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEE
                     HQGLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQD
                     IALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1667..1686)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2277..2296
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     promoter        2441..2652
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             2669..2937
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     regulatory      2983..2992
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2989..2991,2992..2994,2995..3708)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2989..2991,2992..2994,2995..3705)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3034..3055)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3295..3314)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3642..3663
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3736..3759
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     LTR             3822..4336
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    4498..4514
                     /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(join(4522..4528,4529..4546,4547..4552))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4567..4588
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4705..4722)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4876..5464)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4956..4975)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5635..6426,6427..6495))
                     /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"
     primer_bind     6258..6277
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6496..6600)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6668..6686
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6724..6746)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6846..6865
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7059..7081
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7209..7228
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag gcgattagtc
       61 caatttgtta aagacaggat atcagtggtc caggctctag ttttgactca acaatatcac
      121 cagctgaagc ctatagagta cgagccatag ataaaataaa agattttatt tagtctccag
      181 aaaaaggggg gaatgaaaga ccccacctgt aggtttggca agctagctta agtaacgcca
      241 ttttgcaagg catggaaaat acataactga gaatagagaa gttcagatca aggttaggaa
      301 cagagagaca gcagaatatg ggccaaacag gatatctgtg gtaagcagtt cctgccccgg
      361 ctcagggcca agaacagatg gtccccagat gcggtcccgc cctcagcagt ttctagagaa
      421 ccatcagatg tttccagggt gccccaagga cctgaaatga ccctgtgcct tatttgaact
      481 aaccaatcag ttcgcttctc gcttctgttc gcgcgcttct gctccccgag ctcaataaaa
      541 gagcccacaa cccctcactc ggcgcgccag tcctccgata gactgcgtcg cccgggtacc
      601 cgtattccca ataaagcctc ttgctgtttg catccgaatc gtggactcgc tgatccttgg
      661 gagggtctcc tcagattgat tgactgccca cctcgggggt ctttcatttg gaggttccac
      721 cgagatttgg agacccctgc ccagggacca ccgacccccc cgccgggagg taagctggcc
      781 agcggtcgtt tcgtgtctgt ctctgtcttt gtgcgtgttt gtgccggcat ctaatgtttg
      841 cgcctgcgtc tgtactagtt agctaactag ctctgtatct ggcggacccg tggtggaact
      901 gacgagttct gaacacccgg ccgcaaccct gggagacgtc ccagggactt tgggggccgt
      961 ttttgtggcc cgacctgagg aagggagtcg atgtggaatc cgaccccgtc aggatatgtg
     1021 gttctggtag gagacgagaa cctaaaacag ttcccgcctc cgtctgaatt tttgctttcg
     1081 gtttggaacc gaagccgcgc gtcttgtctg ctgcagcgct gcagcatcgt tctgtgttgt
     1141 ctctgtctga ctgtgtttct gtatttgtct gaaaattagg gccagactgt taccactccc
     1201 ttaagtttga ccttaggtca ctggaaagat gtcgagcgga tcgctcacaa ccagtcggta
     1261 gatgtcaaga agagacgttg ggttaccttc tgctctgcag aatggccaac ctttaacgtc
     1321 ggatggccgc gagacggcac ctttaaccga gacctcatca cccaggttaa gatcaaggtc
     1381 ttttcacctg gcccgcatgg acacccagac caggtcccct acatcgtgac ctgggaagcc
     1441 ttggcttttg acccccctcc ctgggtcaag ccctttgtac accctaagcc tccgcctcct
     1501 cttcctccat ccgccccgtc tctccccctt gaacctcctc gttcgacccc gcctcgatcc
     1561 tccctttatc cagccctcac tccttctcta ggcgccggaa ttcaccatgg ccatgattga
     1621 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     1681 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     1741 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
     1801 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     1861 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     1921 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     1981 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     2041 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     2101 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
     2161 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     2221 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     2281 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     2341 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     2401 cttctgagtt taaacatgca gcgctccggt gcccgtcagt gggcagagcg cacatcgccc
     2461 acagtccccg agaagttggg gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg
     2521 cgcggggtaa actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg
     2581 ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc
     2641 gccagaacac agctgaagct agcttcgagg ggctcgcatc tctccttcac gcgcccgccg
     2701 ccctacctga ggccgccatc cacgccggtt gagtcgcgtt ctgccgcctc ccgcctgtgg
     2761 tgcctcctga actgcgtccg ccgtctaggt aagtttaaag ctcaggtcga gaccgggcct
     2821 ttgtccggcg ctcccttgga gcctacctag actcagccgg ctctccacgc tttgcctgac
     2881 cctgcttgct caactctacg tctttgtttc gttttctgtt ctgcgccgtt acagatccaa
     2941 gctgtgaccg gcgcctacat cgatccgcgg agatctaagc ttgccaccat ggtgagcaag
     3001 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     3061 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     3121 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     3181 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
     3241 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     3301 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     3361 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     3421 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     3481 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     3541 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     3601 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     3661 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtaagg atccacgcgt
     3721 gggcccggag gcggcgatta caaggatgac gacgataaga tatgaattct taattaacga
     3781 taaaataaaa gattttattt agtctccaga aaaagggggg aatgaaagac cccacctgta
     3841 ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc atggaaaata cataactgag
     3901 aatagagaag ttcagatcaa ggttaggaac agagagacag cagaatatgg gccaaacagg
     3961 atatctgtgg taagcagttc ctgccccggc tcagggccaa gaacagatgg tccccagatg
     4021 cggtcccgcc ctcagcagtt tctagagaac catcagatgt ttccagggtg ccccaaggac
     4081 ctgaaatgac cctgtgcctt atttgaacta accaatcagt tcgcttctcg cttctgttcg
     4141 cgcgcttctg ctccccgagc tcaataaaag agcccacaac ccctcactcg gcgcgccagt
     4201 cctccgatag actgcgtcgc ccgggtaccc gtgtatccaa taaaccctct tgcagttgca
     4261 tccgacttgt ggtctcgctg ttccttggga gggtctcctc tgagtgattg actacccgtc
     4321 agcgggggtc tttcatgggt aacagtttct tgaagttgga gaacaacatt ctgagggtag
     4381 gagtcgaata ttaagtaatc ctgactcaat tagccactgt tttgaatcca catactccaa
     4441 tactcctgaa atagttcatt atggacagcg cagaaagagc tggggagaat tgtgaaattg
     4501 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     4561 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4621 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4681 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4741 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4801 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     4861 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     4921 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     4981 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     5041 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     5101 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     5161 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     5221 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     5281 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     5341 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     5401 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5461 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5521 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5581 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5641 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5701 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     5761 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     5821 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     5881 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     5941 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     6001 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     6061 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     6121 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     6181 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     6241 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     6301 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     6361 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     6421 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6481 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6541 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6601 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     6661 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     6721 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     6781 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
     6841 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
     6901 cagatgcgta aggagaaaat accgcatcag gcgccattcg ccattcaggc tgcgcaactg
     6961 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
     7021 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     7081 gacggcgcaa ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg
     7141 cctgccacca tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct
     7201 tccccatcgg tgatgtcggc gatataggcg ccagca
//
