LOCUS       sequence-225218-        7165 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sfGFP expressed from hypoxia-inducible minimal CMV promoter and 
            dsRed expressed from consitutive CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7165)
  AUTHORS   Ede C, Chen X, Lin MY, Chen YY
  TITLE     Quantitative Analyses of Core Promoters Enable Precise Engineering 
            of Regulated Gene Expression in Mammalian Cells.
  JOURNAL   ACS Synth Biol. 2016 May 20;5(5):395-404. doi: 
            10.1021/acssynbio.5b00266. Epub 2016 Mar 1.
  PUBMED    26883397
REFERENCE   2  (bases 1 to 7165)
  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: 
            "10.1021/acssynbio.5b00266"; journalName: "ACS Synth Biol"; date: 
            "2016-05-20- 20"; volume: "5"; issue: "5"; pages: "395-404"
COMMENT     Sequence Label: 4xHRE_minCMV-sfGFP-CMV_dsRed Exp
FEATURES             Location/Qualifiers
     source          1..7165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     84..103
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        84..102
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      123..132
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             129..806
                     /codon_start=1
                     /product="rapidly maturing tetrameric variant of DsRed 
                     fluorescent protein (Bevis and Glick, 2002)"
                     /label=DsRed-Express
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGSFIYKVKFIGVNFPSDGPVMQKKTMGWEASTERLYPRDGVLKGEIHK
                     ALKLKDGGHYLVEFKSIYMAKKPVQLPGYYYVDSKLDITSHNEDYTIVEQYERAEGRHH
                     LFL"
     primer_bind     complement(259..277)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     527..546
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     721..744
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    838..1062
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1108..1536
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1195..1214)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1405..1426
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1545..1565)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1550..1879
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1730..1865
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1792..1811
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1946..2740
                     /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2000..2019)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2610..2629
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2914..3035
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2951..2970)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3005..3024
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3084..3100)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3084..3100)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3097..3119)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3108..3124
                     /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(3132..3138,3139..3156,3157..3162))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3177..3198
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3315..3332)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3486..4074)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3566..3585)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4245..5036,5037..5105))
                     /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     4868..4887
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5106..5210)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5457..5495
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5492..5516
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     polyA_signal    6294..6518
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     enhancer        6621..7000
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        join(7001..7165,1..39)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
ORIGIN
        1 ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc
       61 actgcttact ggcttatcga aattaatacg actcactata gggagaccca agctggctag
      121 ccgccaccat ggcctcctcc gaggacgtca tcaaggagtt catgcgcttc aaggtgcgca
      181 tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct
      241 acgagggcac ccagaccgcc aagctgaagg tgaccaaggg cggccccctg cccttcgcct
      301 gggacatcct gtccccccag ttccagtacg gctccaaggt gtacgtgaag caccccgccg
      361 acatccccga ctacaagaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga
      421 acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggctcct
      481 tcatctacaa ggtgaagttc atcggcgtga acttcccctc cgacggcccc gtaatgcaga
      541 agaagactat gggctgggag gcctccaccg agcgcctgta cccccgcgac ggcgtgctga
      601 agggcgagat ccacaaggcc ctgaagctga aggacggcgg ccactacctg gtggagttca
      661 agtccatcta catggccaag aagcccgtgc agctgcccgg ctactactac gtggactcca
      721 agctggacat cacctcccac aacgaggact acaccatcgt ggagcagtac gagcgcgccg
      781 agggccgcca ccacctgttc ctgtagacgg ctgtttaaac ccgctgatca gcctcgactg
      841 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
      901 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
      961 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     1021 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     1081 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
     1141 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     1201 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     1261 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     1321 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     1381 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     1441 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     1501 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt
     1561 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     1621 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     1681 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     1741 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     1801 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg
     1861 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca
     1921 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     1981 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     2041 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     2101 cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac
     2161 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     2221 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     2281 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     2341 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     2401 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     2461 caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     2521 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     2581 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     2641 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     2701 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa
     2761 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     2821 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     2881 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     2941 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     3001 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     3061 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     3121 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     3181 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     3241 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     3301 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     3361 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     3421 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     3481 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     3541 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     3601 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     3661 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     3721 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     3781 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     3841 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     3901 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     3961 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     4021 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     4081 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     4141 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     4201 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     4261 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     4321 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     4381 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     4441 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     4501 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     4561 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     4621 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     4681 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     4741 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     4801 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     4861 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     4921 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     4981 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     5041 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     5101 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     5161 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     5221 cgaaaagtgc cacctgacgt cgacggatcg ggagatcttt gtatttaatt aagaccttga
     5281 gtacgtgcgt ctctgcacgt atggcgatta agaccttgag tacgtgcgtc tctgcacgta
     5341 tggcgattaa gaccttgagt acgtgcgtct ctgcacgtat ggcgattaag accttgagta
     5401 cgtgcgtctc tgcacgtatg gcggatcttc gcgattacgc aataatggta ccatctggta
     5461 ggcgtgtacg gtgggaggtc tatataagca gagctcgttt agtgaaccgt cagatcatct
     5521 agcgccgcca ccatgagtaa aggtgaagaa ctgttcaccg gtgttgttcc gatcctggtt
     5581 gaactggatg gtgatgttaa cggccacaaa ttctctgttc gtggtgaagg tgaaggtgat
     5641 gcaaccaacg gtaaactgac cctgaaattc atctgcacta ccggtaaact gccggttcca
     5701 tggccgactc tggtgactac cctgacctat ggtgttcagt gtttttctcg ttacccggat
     5761 cacatgaagc agcatgattt cttcaaatct gcaatgccgg aaggttatgt acaggagcgc
     5821 accatttctt tcaaagacga tggcacctac aaaacccgtg cagaggttaa atttgaaggt
     5881 gatactctgg tgaaccgtat tgaactgaaa ggcattgatt tcaaagagga cggcaacatc
     5941 ctgggccaca aactggaata taacttcaac tcccataacg tttacatcac cgcagacaaa
     6001 cagaagaacg gtatcaaagc taacttcaaa attcgccata acgttgaaga cggtagcgta
     6061 cagctggcgg accactacca gcagaacact ccgatcggtg atggtccggt tctgctgccg
     6121 gataaccact acctgtccac ccagtctaaa ctgtccaaag acccgaacga aaagcgcgac
     6181 cacatggtgc tgctggagtt cgttactgca gcaggtatca cgcacggcat ggatgaactc
     6241 tacaaataag cggccgccta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc
     6301 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     6361 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     6421 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     6481 caatagcagg catgctgggg atgcggtggg ctctatggcc gcggaattag cctatgtttt
     6541 ttggaacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt
     6601 acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac
     6661 ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg
     6721 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
     6781 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
     6841 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa
     6901 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac
     6961 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta
     7021 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga
     7081 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa
     7141 ctccgcccca ttgacgcaaa tgggc
//
