LOCUS       sequence-401528-        8302 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to Fc(DAPA)-Avi-6xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8302)
  AUTHORS   Wojtowicz WM, Vielmetter J, Fernandes RA, Siepe DH, Eastman CL, 
            Chisholm GB, Cox S, Klock H, Anderson PW, Rue SM, Miller JJ, Glaser 
            SM, Bragstad ML, Vance J, Lam AW, Lesley SA, Zinn K, Garcia KC
  TITLE     A Human IgSF Cell-Surface Interactome Reveals a Complex Network of 
            Protein-Protein Interactions.
  JOURNAL   Cell. 2020 Aug 20;182(4):1027-1043.e17. doi: 
            10.1016/j.cell.2020.07.025.
  PUBMED    32822567
REFERENCE   2  (bases 1 to 8302)
  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: 
            "10.1016/j.cell.2020.07"; journalName: "Cell"; date: "2020-08-20- 
            20"; volume: "182"; issue: "4"; pages: "1027-1043.e17"
COMMENT     Sequence Label: pD649-HAsp-FLT1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8302
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        77..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        815..833
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     815..832
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     846..865
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             3878..3922
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3923..3940
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4080..4097)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4086..4197
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4315..4335)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4320..4649
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4500..4635
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4562..4581
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4697..4744
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4763..5362
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4763..4782)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5259..5279
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5492..5613
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5529..5548)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5583..5602
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5666..6457,6458..6526))
                     /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     6289..6308
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6527..6618)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6660..6679
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6685..6704
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6790..7378
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7279..7298
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7532..7549
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7647..7664
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7648..7664
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8120..8136
                     /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)."
ORIGIN
        1 ggcattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat
      661 ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc aagagtgacg
      721 taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa cgcggctaca
      781 attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat agaataacat
      841 ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc tcggttctat
      901 cgaaaacgcg cctgctagcg ccaccatgaa gaccatcatt gccctgagct acatcttctg
      961 cctggtgttc gctgcggccg cctctaagct gaaagatcct gagctgagcc tgaagggcac
     1021 ccagcatatt atgcaggccg gacagaccct gcatctgcag tgtagaggcg aagccgctca
     1081 caagtggtcc ctgcctgaga tggtgtccaa agagagcgag cggctgagca tcacaaagag
     1141 cgcctgcggc agaaacggca agcagttctg tagcaccctg acactgaaca cagcccaggc
     1201 caatcacacc ggcttctaca gctgcaagta cctggccgtg cctaccagca agaagaaaga
     1261 aaccgagagc gccatctaca tcttcatcag cgacaccggc agacccttcg tggaaatgta
     1321 cagcgagatc cccgagatca tccacatgac cgagggcaga gagctggtca tcccttgcag
     1381 agtgacaagc cccaacatca cagtgaccct gaagaagttc cctctggaca cactgatccc
     1441 cgacggcaag agaatcatct gggacagccg gaagggcttc atcatcagca acgccaccta
     1501 caaagagatc ggcctgctga catgcgaggc cacagtgaat ggccacctgt acaagaccaa
     1561 ctatctgacc cacagacaga ccaacaccat catcgacgtg cagatcagca cccctagacc
     1621 tgtgaagctg ctgagaggac acaccctggt gctgaattgc accgccacca cacctctgaa
     1681 caccagagtg cagatgacct ggtcttaccc cgacgagaag aacaagagag ccagcgtcag
     1741 aaggcggatc gaccagagca atagccacgc caacatcttc tactccgtgc tgaccatcga
     1801 caagatgcag aacaaggaca agggcctgta cacctgtaga gtgcggagcg gccctagctt
     1861 caagtccgtg aataccagcg tgcacatcta cgacaaggcc ttcatcaccg tgaagcaccg
     1921 gaagcagcag gtcctggaaa cagtggccgg caagaggtcc tacagactga gcatgaaggt
     1981 caaagccttt ccatctcctg aggtcgtgtg gctgaaggat ggactgcctg ccacagagaa
     2041 gtccgccaga tacctgacaa gaggctacag cctgatcatc aaggacgtga ccgaagagga
     2101 cgccggcaac tacaccatcc tgctgtccat caagcagagc aacgtgttca agaacctgac
     2161 cgccactctg atcgtgaacg tgaagcccca gatctatgag aaggccgtgt ccagctttcc
     2221 cgatcctgct ctgtatcctc tgggcagcag acagatcctg acctgcacag cctatggcat
     2281 ccctcagcct accatcaagt ggttctggca cccctgcaac cacaaccaca gcgaggccag
     2341 atgcgacttc tgcagcaaca acgaggaaag cttcatcctg gacgccgaca gcaacatggg
     2401 caacagaatc gagagcatca cccagcggat ggccattatc gagggcaaga acaaaatggc
     2461 cagcacactg gtggtggccg actccagaat cagcggcatc tatatctgta tcgccagcaa
     2521 caaagtgggc accgtgggca gaaacatcag cttctacatc accgatgtgc ccaacggctt
     2581 ccacgtgaac ctggaaaaga tgcccaccga aggcgaggac ctgaagctga gctgcaccgt
     2641 gaacaagttt ctgtaccgcg acgtgacctg gattctgctg cggacagtga acaaccggac
     2701 catgcactac agcatcagca agcagaagat ggctatcacc aaagagcaca gcatcacact
     2761 gaacctcacc atcatgaacg tgtcactgca ggacagcggc acctacgcct gtagagccag
     2821 aaatgtgtac accggcgagg aaatcctgca gaaaaaagag atcaccatcc gggaccaaga
     2881 ggccccttac ctgctgagaa atctgagcga tcacaccgtg gccatcagca gcagcacaac
     2941 actggattgc catgccaatg gcgtgcccga gcctcagatc acctggttta agaacaacca
     3001 caagatccag caagagcccg gcatcatcct cggacctggc agctctaccc tgttcatcga
     3061 gagagtcact gaagaggacg agggcgtgta ccactgcaag gccacaaatc agaaaggcag
     3121 cgtggaaagc agcgcctacc tgacagtgca gggcaccagc gacaagagca acctggaagg
     3181 cgcgccgggc ggcgacaaga cccacacctg ccccccctgc ccagctccag aactgctggg
     3241 agggccttcc gtgttcctgt tcccccccaa gcccaaggac accctgatga tcagcaggac
     3301 ccccgaggtg acctgcgtgg tggtggccgt gtcccacgag gacccagagg tgaagttcaa
     3361 ctggtacgtg gacggcgtgg aggtgcacaa cgccaagacc aagcccagag aggagcagta
     3421 caacagcacc tacagggtgg tgtccgtgct gaccgtgctg caccaggact ggctgaacgg
     3481 caaagaatac aagtgcaaag tctccaacaa ggccctggct gccccaatcg aaaagacaat
     3541 cagcaaggcc aagggccagc cacgggagcc ccaggtgtac accctgcccc ccagccggga
     3601 ggagatgacc aagaaccagg tgtccctgac ctgtctggtg aagggcttct accccagcga
     3661 tatcgccgtg gagtgggaga gcaacggcca gcccgagaac aactacaaga ccaccccccc
     3721 agtgctggac agcgacggca gcttcttcct gtacagcaag ctgaccgtgg acaagtccag
     3781 gtggcagcag ggcaacgtgt tcagctgcag cgtgatgcac gaggccctgc acaaccacta
     3841 cacccagaag tccctgagcc tgagccccgg cggcagcggt cttaatgata ttttcgaggc
     3901 tcagaagatt gagtggcatg aacaccacca ccatcaccac taaggccggc ctgtagtcac
     3961 tggcacaacg cgttacccgg taagccaatc gggtatacac ggtcgtcata ctgcagacag
     4021 ggttcttcta ctttgcaaga tagtcttgag tagtaaaata aatagataga gaaaatcagc
     4081 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     4141 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcaca
     4201 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     4261 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg
     4321 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     4381 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     4441 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     4501 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     4561 ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag
     4621 gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg
     4681 gatctgatca gcacgtgttg acaattaatc atcggcatag tatatcggca tagtataata
     4741 cgacaaggtg aggaactaaa ccatgaccga gtacaagccc acggtgcgcc tcgccacccg
     4801 cgacgacgtc cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac
     4861 gcgccacacc gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt
     4921 cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt
     4981 ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc
     5041 gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct
     5101 ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga
     5161 ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg
     5221 cgccggggtg cccgccttcc tggagacatc cgcgccccgc aacctcccct tctacgagcg
     5281 gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat
     5341 gacccgcaag cccggtgcct gacacgtgct acgagatttc gattccaccg ccgccttcta
     5401 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     5461 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     5521 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     5581 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     5641 ctagagcttg gcgtaatcat ggtcattacc aatgcttaat cagtgaggca cctatctcag
     5701 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     5761 tacgggaggg cttaccatct ggccccagcg ctgcgatgat accgcgagaa ccacgctcac
     5821 cggctccgga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     5881 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     5941 gttcgccagt taatagtttg cgcaacgttg ttgccatcgc tacaggcatc gtggtgtcac
     6001 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6061 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6121 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6181 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     6241 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     6301 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     6361 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     6421 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     6481 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatattc ttcctttttc
     6541 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     6601 tttagaaaaa taaacaaata ggggtcagtg ttacaaccaa ttaaccaatt ctgaacatta
     6661 tcgcgagccc atttatacct gaatatggct cataacaccc cttgctcatg accaaaatcc
     6721 cttaacgtga gttacgcgcg cgtcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     6781 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     6841 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     6901 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagc
     6961 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     7021 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     7081 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     7141 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     7201 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     7261 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     7321 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     7381 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     7441 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     7501 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaagg
     7561 cgagagtagg gaactgccag gcatcaaact aagcagaagg cccctgacgg atggcctttt
     7621 tgcgtttcta caaactcttt ctgtgttgta aaacgacggc cagtcttaag ctcgggcccc
     7681 ctgggcggtt ctgataacga gtaatcgtta atccgcaaat aacgtaaaaa cccgcttcgg
     7741 cgggtttttt tatgggggga gtttagggaa agagcatttg tcagaatatt taagggcgcc
     7801 tgtcactttg cttgatatat gagaattatt taaccttata aatgagaaaa aagcaacgca
     7861 ctttaaataa gatacgttgc tttttcgatt gatgaacacc tataattaaa ctattcatct
     7921 attatttatg attttttgta tatacaatat ttctagtttg ttaaagagaa ttaagaaaat
     7981 aaatctcgaa aataataaag ggaaaatcag tttttgatat caaaattata catgtcaacg
     8041 ataatacaaa atataataca aactataaga tgttatcagt atttattatc atttagaata
     8101 aattttgtgt cgcccttaat tgtgagcgga taacaattac gagcttcatg cacagtggcg
     8161 ttgactcaat attggccatt agccatatta ttcattggtt atatagcata aatcaatatt
     8221 ggctattggc cattgcatac gttgtatcta tatcataata tgtacattta tattggctca
     8281 tgtccaatat gaccgccatg tt
//
