LOCUS       sequence-211329-        7519 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  positive control  for GFP expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7519)
  AUTHORS   Wei W, Tweardy DJ, Zhang M, Zhang X, Landua J, Petrovic I, Bu W, 
            Roarty K, Hilsenbeck SG, Rosen JM, Lewis MT
  TITLE     STAT3 signaling is activated preferentially in tumor-initiating 
            cells in claudin-low models of human breast cancer.
  JOURNAL   Stem Cells. 2014 Oct;32(10):2571-82. doi: 10.1002/stem.1752.
  PUBMED    24891218
REFERENCE   2  (bases 1 to 7519)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1002/stem"; 
            journalName: "Stem Cells"; date: "2014-10"; volume: "32"; issue: 
            "10"; pages: "2571-82"
COMMENT     Sequence Label: EFS-GFP
FEATURES             Location/Qualifiers
     source          1..7519
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     19..35
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     20..36
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(78..111)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        166..377
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      451..460
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(457..459,460..462,463..1176)
                     /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(457..459,460..462,463..1173)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(502..523)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(763..782)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1110..1131
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1201..1789
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1254..1274)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1672..1683)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             1876..2109
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2181..2302
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2218..2237)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2272..2291
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2342..2477
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2497..2516)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2498..2516)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2526..2543)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2526..2542)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2535..2557)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2684..3139
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2771..2790)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2981..3002
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3165..3269
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3270..3338,3339..4130)
                     /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     complement(3488..3507)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4301..4889
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4790..4809
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5043..5060
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5177..5198
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5213..5218,5219..5236,5237..5243)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5251..5267
                     /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)."
     primer_bind     5256..5278
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5275..5291
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5275..5291
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5310..5330
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5312..5330
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5358..5584
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5585..5765
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5812..5937
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6430..6663
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6848..6892
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    7159..7276
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_feature    7363..7480
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 gtaccgggcc cgctctagtc gaggtcgacg gtatcgataa gctcgcttca cgagattcca
       61 gcaggtcgag ggacctaata acttcgtata gcatacatta tacgaagtta tattaagggt
      121 tccaagctta agcggccgca tcgattggct ccggtgcccg tcagtgggca gagcgcacat
      181 cgcccacagt ccccgagaag ttggggggag gggtcggcaa ttgaaccggt gcctagagaa
      241 ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt tttcccgagg
      301 gtgggggaga accgtatata agtgcagtag tcgccgtgaa cgttcttttt cgcaacgggt
      361 ttgccgccag aacacaggtg tcgtgacgcg ggatccggta ctcgagatct gcgatctaag
      421 taagcttggc attccggtac tgttggtaaa gccaccatgg tgagcaaggg cgaggagctg
      481 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
      541 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
      601 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc
      661 gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc
      721 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
      781 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
      841 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc
      901 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc
      961 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc
     1021 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg
     1081 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     1141 gggatcactc tcggcatgga cgagctgtac aagtaaagcg gccgcgactc tagagtcgac
     1201 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     1261 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     1321 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
     1381 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
     1441 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
     1501 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     1561 ttgggcactg acaattccgt ggtgttgtcg gggaagctga cgtcctttcc atggctgctc
     1621 gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
     1681 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
     1741 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcc tggaattcga
     1801 gctcggtacc tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa
     1861 aagaaaaggg gggactggaa gggctaattc actcccaacg aagacaagat ctgctttttg
     1921 cttgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     1981 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     2041 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     2101 tctctagcag tagtagttca tgtcatctta ttattcagta tttataactt gcaaagaaat
     2161 gaatatcaga gagtgagagg aacttgttta ttgcagctta taatggttac aaataaagca
     2221 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     2281 ccaaactcat caatgtatct tatcatgtct ggctctagct atcccgcccc taactccgcc
     2341 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     2401 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     2461 aggctttttt ggaggcctag ggacgtaccc aattcgccct atagtgagtc gtattacgcg
     2521 cgctcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
     2581 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
     2641 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc
     2701 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     2761 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     2821 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     2881 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     2941 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     3001 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     3061 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     3121 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     3181 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     3241 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     3301 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     3361 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     3421 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     3481 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     3541 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     3601 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     3661 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     3721 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     3781 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     3841 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     3901 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     3961 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     4021 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     4081 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     4141 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     4201 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     4261 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     4321 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4381 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     4441 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     4501 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     4561 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     4621 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     4681 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     4741 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     4801 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     4861 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     4921 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     4981 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     5041 gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg
     5101 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     5161 gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact
     5221 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     5281 acagctatga ccatgattac gccaagcgcg caattaaccc tcactaaagg gaacaaaagc
     5341 tggagctgca agcttaatgt agtcttatgc aatactcttg tagtcttgca acatggtaac
     5401 gatgagttag caacatgcct tacaaggaga gaaaaagcac cgtgcatgcc gattggtgga
     5461 agtaaggtgg tacgatcgtg ccttattagg aaggcaacag acgggtctga catggattgg
     5521 acgaaccact gaattgccgc attgcagaga tattgtattt aagtgcctag ctcgatacat
     5581 aaacgggtct ctctggttag accagatctg agcctgggag ctctctggct aactagggaa
     5641 cccactgctt aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct
     5701 gttgtgtgac tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc
     5761 tagcagtggc gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga
     5821 cgcaggactc ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt
     5881 acgccaaaaa ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt
     5941 attaagcggg ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag
     6001 aaaaaatata aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt
     6061 aatcctggcc tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca
     6121 tcccttcaga caggatcaga agaacttaga tcattatata atacagtagc aaccctctat
     6181 tgtgtgcatc aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa
     6241 gagcaaaaca aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg
     6301 aggagatatg agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga
     6361 accattagga gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc
     6421 agtgggaata ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc
     6481 agcgtcaatg acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca
     6541 gaacaatttg ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg
     6601 catcaagcag ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct
     6661 cctggggatt tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc
     6721 tagttggagt aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga
     6781 cagagaaatt aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca
     6841 gcaagaaaag aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg
     6901 gtttaacata acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt
     6961 ggtaggttta agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata
     7021 ttcaccatta tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg
     7081 aatagaagaa gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc
     7141 tcgacggtat cggttaactt ttaaaagaaa aggggggatt ggggggtaca gtgcagggga
     7201 aagaatagta gacataatag caacagacat acaaactaaa gaattacaaa aacaaattac
     7261 aaaaattcaa aattttatcg atcacgagac tagcctccta gcccgggctc gagcagctga
     7321 agcttgcatg ccctgcagac aaatggcagt attcatccac aattttaaaa gaaaaggggg
     7381 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
     7441 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag
     7501 cagagatcca gtttggtta
//
