LOCUS       sequence-460790-        8180 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty backbone for expression of Ovalbumin with Bi-cistronic 
            expression ZsGreen.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8180)
  AUTHORS   Hanggi K, Li J, Gangadharan A, Liu X, Celias DP, Osunmakinde O, 
            Keske A, Davis J, Ahmad F, Giron A, Anadon CM, Gardner A, DeNardo 
            DG, Shaw TI, Beg AA, Yu X, Ruffell B
  TITLE     Interleukin-1alpha release during necrotic-like cell death generates
            myeloid-driven immunosuppression that restricts anti-tumor immunity.
  JOURNAL   Cancer Cell. 2024 Nov 13:S1535-6108(24)00402-1. doi: 
            10.1016/j.ccell.2024.10.014.
  PUBMED    39577420
REFERENCE   2  (bases 1 to 8180)
  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"; journalName: "Cancer 
            Cell. 2024 Nov 13:S1535-6108(24)00402-1. doi: 
            10.1016/j.ccell.2024.10.014."
COMMENT     Sequence Label: pHIV-EF1a-2A-Ovalbumin(dN)-ZsGreen
FEATURES             Location/Qualifiers
     source          1..8180
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(554..570)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(555..571)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             1080..1260
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(1322..1910)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1402..1421)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2081..2872,2873..2941))
                     /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     2704..2723
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2942..3046)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3121..3140)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3312..3691
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3693..3891
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3846..3866
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             3909..4089
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    4136..4261
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    4758..4991
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             5176..5220
                     /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    5518..5635
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        5698..6876
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          5929..6867
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     6824..6844
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             7566..7589
                     /codon_start=1
                     /product="antigenic CD8+ T cell epitope from chicken 
                     ovalbumin"
                     /label=OVA peptide
                     /translation="SIINFEKL"
     CDS             7959..8012
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             join(8025..8180,1..540)
                     /codon_start=1
                     /product="Zoanthus green fluorescent protein"
                     /label=ZsGreen1
                     /note="mammalian codon-optimized"
                     /translation="MAQSKHGLTKEMTMKYRMEGCVDGHKFVITGEGIGYPFKGKQAIN
                     LCVVEGGPLPFAEDILSAAFMYGNRVFTEYPQDIVDYFKNSCPAGYTWDRSFLFEDGAV
                     CICNADITVSVEENCMYHESKFYGVNFPADGPVMKKMTDNWEPSCEKIIPVPKQGILKG
                     DVSMYLLLKDGGRLRCQFDTVYKAKSVPRKMPDWHFIQHKLTREDRSDAKNQKWHLTEH
                     AIASGSALP"
ORIGIN
        1 cccttgccct tcgccgagga catcttgtcc gccgccttca tgtacggcaa ccgcgtgttc
       61 accgagtacc cccaggacat cgtcgactac ttcaagaact cctgccccgc cggctacacc
      121 tgggaccgct ccttcctgtt cgaggacggc gccgtgtgca tctgcaacgc cgacatcacc
      181 gtgagcgtgg aggagaactg catgtaccac gagtccaagt tctacggcgt gaacttcccc
      241 gccgacggcc ccgtgatgaa gaagatgacc gacaactggg agccctcctg cgagaagatc
      301 atccccgtgc ccaagcaggg catcttgaag ggcgacgtga gcatgtacct gctgctgaag
      361 gacggtggcc gcttgcgctg ccagttcgac accgtgtaca aggccaagtc cgtgccccgc
      421 aagatgcccg actggcactt catccagcac aagctgaccc gcgaggaccg cagcgacgcc
      481 aagaaccaga agtggcacct gaccgagcac gccatcgcct ccggctccgc cttgccctaa
      541 ggatcggccg catcgatacc gtcgacctcg atcgagacct agaaaaacat ggagcaatca
      601 caagtagcaa tacagcagct accaatgctg attgtgcctg gctagaagca caagaggagg
      661 aggaggtggg ttttccagtc acacctcagg tacctttaag accaatgact tacaaggcag
      721 ctgtagatct tagccacttt ttaaaagaaa aggggggact ggaagggcta attcactccc
      781 aacgaagaca agatatcctt gatctgtgga tctaccacac acaaggctac ttccctgatt
      841 ggcagaacta cacaccaggg ccagggatca gatatccact gacctttgga tggtgctaca
      901 agctagtacc agttgagcaa gagaaggtag aagaagccaa tgaaggagag aacacccgct
      961 tgttacaccc tgtgagcctg catgggatgg atgacccgga gagagaagta ttagagtgga
     1021 ggtttgacag ccgcctagca tttcatcaca tggcccgaga gctgcatccg gactgtactg
     1081 ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac
     1141 tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt
     1201 gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca
     1261 gcatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     1321 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     1381 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     1441 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     1501 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     1561 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     1621 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     1681 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     1741 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     1801 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     1861 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     1921 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     1981 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     2041 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     2101 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     2161 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     2221 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     2281 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     2341 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     2401 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     2461 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     2521 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     2581 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     2641 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     2701 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     2761 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     2821 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     2881 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     2941 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     3001 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     3061 aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag
     3121 tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga
     3181 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa
     3241 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag
     3301 atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
     3361 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
     3421 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
     3481 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
     3541 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
     3601 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
     3661 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
     3721 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
     3781 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
     3841 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gcgcgttttg
     3901 cctgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     3961 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     4021 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     4081 tctctagcag tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc
     4141 tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt
     4201 gagtacgcca aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt
     4261 cagtattaag cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg
     4321 aaagaaaaaa tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc
     4381 agttaatcct ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca
     4441 accatccctt cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct
     4501 ctattgtgtg catcaaagga tagagataaa agacaccaag gaagctttag acaagataga
     4561 ggaagagcaa aacaaaagta agaccaccgc acagcaagcg gccggccgct gatcttcaga
     4621 cctggaggag gagatatgag ggacaattgg agaagtgaat tatataaata taaagtagta
     4681 aaaattgaac cattaggagt agcacccacc aaggcaaaga gaagagtggt gcagagagaa
     4741 aaaagagcag tgggaatagg agctttgttc cttgggttct tgggagcagc aggaagcact
     4801 atgggcgcag cgtcaatgac gctgacggta caggccagac aattattgtc tggtatagtg
     4861 cagcagcaga acaatttgct gagggctatt gaggcgcaac agcatctgtt gcaactcaca
     4921 gtctggggca tcaagcagct ccaggcaaga atcctggctg tggaaagata cctaaaggat
     4981 caacagctcc tggggatttg gggttgctct ggaaaactca tttgcaccac tgctgtgcct
     5041 tggaatgcta gttggagtaa taaatctctg gaacagattt ggaatcacac gacctggatg
     5101 gagtgggaca gagaaattaa caattacaca agcttaatac actccttaat tgaagaatcg
     5161 caaaaccagc aagaaaagaa tgaacaagaa ttattggaat tagataaatg ggcaagtttg
     5221 tggaattggt ttaacataac aaattggctg tggtatataa aattattcat aatgatagta
     5281 ggaggcttgg taggtttaag aatagttttt gctgtacttt ctatagtgaa tagagttagg
     5341 cagggatatt caccattatc gtttcagacc cacctcccaa ccccgagggg acccgacagg
     5401 cccgaaggaa tagaagaaga aggtggagag agagacagag acagatccat tcgattagtg
     5461 aacggatcgg cactgcgtgc gccaattctg cagacaaatg gcagtattca tccacaattt
     5521 taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag acataatagc
     5581 aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa attttcgggt
     5641 ttattacagg gacagcagag atccagtttg gttagtaccg ggcccgctct agcgtgaggc
     5701 tccggtgccc gtcagtgggc agagcgcaca tcgcccacag tccccgagaa gttgggggga
     5761 ggggtcggca attgaaccgg tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat
     5821 gtcgtgtact ggctccgcct ttttcccgag ggtgggggag aaccgtatat aagtgcagta
     5881 gtcgccgtga acgttctttt tcgcaacggg tttgccgcca gaacacaggt aagtgccgtg
     5941 tgtggttccc gcgggcctgg cctctttacg ggttatggcc cttgcgtgcc ttgaattact
     6001 tccacctggc tgcagtacgt gattcttgat cccgagcttc gggttggaag tgggtgggag
     6061 agttcgaggc cttgcgctta aggagcccct tcgcctcgtg cttgagttga ggcctggcct
     6121 gggcgctggg gccgccgcgt gcgaatctgg tggcaccttc gcgcctgtct cgctgctttc
     6181 gataagtctc tagccattta aaatttttga tgacctgctg cgacgctttt tttctggcaa
     6241 gatagtcttg taaatgcggg ccaagatctg cacactggta tttcggtttt tggggccgcg
     6301 ggcggcgacg gggcccgtgc gtcccagcgc acatgttcgg cgaggcgggg cctgcgagcg
     6361 cggccaccga gaatcggacg ggggtagtct caagctggcc ggcctgctct ggtgcctggc
     6421 ctcgcgccgc cgtgtatcgc cccgccctgg gcggcaaggc tggcccggtc ggcaccagtt
     6481 gcgtgagcgg aaagatggcc gcttcccggc cctgctgcag ggagctcaaa atggaggacg
     6541 cggcgctcgg gagagcgggc gggtgagtca cccacacaaa ggaaaagggc ctttccgtcc
     6601 tcagccgtcg cttcatgtga ctccacggag taccgggcgc cgtccaggca cctcgattag
     6661 ttctcgagct tttggagtac gtcgtcttta ggttgggggg aggggtttta tgcgatggag
     6721 tttccccaca ctgagtgggt ggagactgaa gttaggccag cttggcactt gatgtaattc
     6781 tccttggaat ttgccctttt tgagtttgga tcttggttca ttctcaagcc tcagacagtg
     6841 gttcaaagtt tttttcttcc atttcaggtg tcgtgagcgg ccgctgaact gaattcatcg
     6901 acgttaacta ttctagagta cccgggctag gatccatgag caccaggaca cagataaata
     6961 aggttgttcg ctttgataaa cttccaggat tcggagacag tattgaagct cagtgtggca
     7021 catctgtaaa cgttcactct tcacttagag acatcctcaa ccaaatcacc aaaccaaatg
     7081 atgtttattc gttcagcctt gccagtagac tttatgctga agagagatac ccaatcctgc
     7141 cagaatactt gcagtgtgtg aaggaactgt atagaggagg cttggaacct atcaactttc
     7201 aaacagctgc agatcaagcc agagagctca tcaattcctg ggtagaaagt cagacaaatg
     7261 gaattatcag aaatgtcctt cagccaagct ccgtggattc tcaaactgca atggttctgg
     7321 ttaatgccat tgtcttcaaa ggactgtggg agaaagcatt taaggatgaa gacacacaag
     7381 caatgccttt cagagtgact gagcaagaaa gcaaacctgt gcagatgatg taccagattg
     7441 gtttatttag agtggcatca atggcttctg agaaaatgaa gatcctggag cttccatttg
     7501 ccagtgggac aatgagcatg ttggtgctgt cgcctgatga agtctcaggc cttgagcagc
     7561 ttgagagtat aatcaacttt gaaaaactga ctgaatggac cagttctaat gttatggaag
     7621 agaggaagat caaagtgtac ttacctcgca tgaagatgga ggaaaaatac aacctcacat
     7681 ctgtcttaat ggctatgggc attactgacg tgtttagctc ttcagccaat ctgtctggca
     7741 tctcctcagc agagagcctg aagatatctc aagctgtcca tgcagcacat gcagaaatca
     7801 atgaagcagg cagagaggtg gtagggtcag cagaggctgg agtggatgct gcaagcgtct
     7861 ctgaagaatt tagggctgac catccattcc tcttctgtat caagcacatc gcaaccaacg
     7921 ccgttctctt ctttggcaga tgtgtttccc ctggatccga gggcagagga agtcttctaa
     7981 catgcggtga cgtggaggag aatcccggcc ctatggccac aaccatggcc cagtccaagc
     8041 acggcctgac caaggagatg accatgaagt accgcatgga gggctgcgtg gacggccaca
     8101 agttcgtgat caccggcgag ggcatcggct accccttcaa gggcaagcag gccatcaacc
     8161 tgtgcgtggt ggagggcggc
//
