LOCUS       sequence-405787-        7372 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 7372)
  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 7372)
  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-LAG3-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..7372
                     /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             2948..2992
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2993..3010
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3150..3167)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3156..3267
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3385..3405)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3390..3719
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3570..3705
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3632..3651
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3767..3814
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3833..4432
                     /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(3833..3852)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4329..4349
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4562..4683
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4599..4618)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4653..4672
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4736..5527,5528..5596))
                     /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     5359..5378
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5597..5688)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5730..5749
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5755..5774
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5860..6448
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6349..6368
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6602..6619
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6717..6734
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6718..6734
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7190..7206
                     /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 ccgttccagt tgtttgggct caagaaggtg cccctgctca
     1021 gctgccttgc agccctacaa tcccactgca ggatctgagc ctgcttcgga gagctggcgt
     1081 gacatggcag caccagcctg attctggacc tcctgctgct gctcctggac atccacttgc
     1141 tccaggacct catcctgccg ctcctagttc ttggggccct agacctagac ggtacaccgt
     1201 gctgtctgtt ggacctggcg gactgagatc tggcagactg cctctgcaac ccagagtgca
     1261 gctggacgaa agaggcagac agaggggcga tttcagcctg tggcttagac ctgccagaag
     1321 ggctgacgcc ggcgagtata gagctgccgt gcacctgaga gatagagccc tgagctgccg
     1381 gctgagactg agactgggac aagccagcat gaccgcctct ccacctggat ctctgagagc
     1441 cagcgattgg gtcatcctga actgcagctt ctcccggcct gatagaccag ccagcgtgca
     1501 ctggttcaga aacagaggcc agggcagagt gcccgtgcgg gaatctcctc atcatcacct
     1561 ggccgagagc tttctgttcc tgcctcaagt gtcccctatg gactctggcc cttggggctg
     1621 tatcctgacc tacagagatg gcttcaacgt gtccatcatg tacaacctga ccgtgctggg
     1681 cctcgagcct cctacacctc tgacagttta tgccggcgct ggcagcagag tgggcctgcc
     1741 ttgtagactt cctgccggcg tgggcaccag aagctttctg acagccaagt ggacacctcc
     1801 tggcggcgga cctgatttgc ttgtgacagg cgacaacggc gacttcaccc tgagactcga
     1861 ggatgtgtct caggcccagg ccggcacata cacctgtcac atccatctgc aagaacagca
     1921 gctgaacgcc accgtgacac tggccatcat cacagtgacc cctaagagct ttggcagccc
     1981 tggcagcctg ggaaagctgc tgtgtgaagt gacacccgtg tccggccaag agcgctttgt
     2041 ttggagcagc ctggataccc ctagccagag aagctttagc ggcccttggc tggaagccca
     2101 agaagcccag ctgctttctc agccttggca gtgccagctg taccagggcg aaagactgct
     2161 gggagccgcc gtgtacttta ccgagcttag ttctcctggc gctcaaagaa gcggcagagc
     2221 acctggtgca ttgcctgccg gacatcttgg cgcgccgggc ggcgacaaga cccacacctg
     2281 ccccccctgc ccagctccag aactgctggg agggccttcc gtgttcctgt tcccccccaa
     2341 gcccaaggac accctgatga tcagcaggac ccccgaggtg acctgcgtgg tggtggccgt
     2401 gtcccacgag gacccagagg tgaagttcaa ctggtacgtg gacggcgtgg aggtgcacaa
     2461 cgccaagacc aagcccagag aggagcagta caacagcacc tacagggtgg tgtccgtgct
     2521 gaccgtgctg caccaggact ggctgaacgg caaagaatac aagtgcaaag tctccaacaa
     2581 ggccctggct gccccaatcg aaaagacaat cagcaaggcc aagggccagc cacgggagcc
     2641 ccaggtgtac accctgcccc ccagccggga ggagatgacc aagaaccagg tgtccctgac
     2701 ctgtctggtg aagggcttct accccagcga tatcgccgtg gagtgggaga gcaacggcca
     2761 gcccgagaac aactacaaga ccaccccccc agtgctggac agcgacggca gcttcttcct
     2821 gtacagcaag ctgaccgtgg acaagtccag gtggcagcag ggcaacgtgt tcagctgcag
     2881 cgtgatgcac gaggccctgc acaaccacta cacccagaag tccctgagcc tgagccccgg
     2941 cggcagcggt cttaatgata ttttcgaggc tcagaagatt gagtggcatg aacaccacca
     3001 ccatcaccac taaggccggc ctgtagtcac tggcacaacg cgttacccgg taagccaatc
     3061 gggtatacac ggtcgtcata ctgcagacag ggttcttcta ctttgcaaga tagtcttgag
     3121 tagtaaaata aatagataga gaaaatcagc ctcgactgtg ccttctagtt gccagccatc
     3181 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     3241 ttcctaataa aatgaggaaa ttgcatcaca acactcaacc ctatctcggt ctattctttt
     3301 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     3361 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     3421 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     3481 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     3541 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     3601 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     3661 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     3721 agctcccggg agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc
     3781 atcggcatag tatatcggca tagtataata cgacaaggtg aggaactaaa ccatgaccga
     3841 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct
     3901 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat
     3961 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
     4021 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     4081 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     4141 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     4201 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
     4261 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacatc
     4321 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     4381 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacacgtgct
     4441 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     4501 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     4561 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     4621 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4681 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcattacc
     4741 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     4801 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagcg
     4861 ctgcgatgat accgcgagaa ccacgctcac cggctccgga tttatcagca ataaaccagc
     4921 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     4981 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5041 ttgccatcgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5101 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5161 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5221 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5281 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5341 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5401 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     5461 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     5521 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     5581 aatgttgaat actcatattc ttcctttttc aatattattg aagcatttat cagggttatt
     5641 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggtcagtg
     5701 ttacaaccaa ttaaccaatt ctgaacatta tcgcgagccc atttatacct gaatatggct
     5761 cataacaccc cttgctcatg accaaaatcc cttaacgtga gttacgcgcg cgtcgttcca
     5821 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     5881 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     5941 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     6001 tactgttctt ctagtgtagc cgtagttagc ccaccacttc aagaactctg tagcaccgcc
     6061 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     6121 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     6181 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     6241 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     6301 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     6361 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     6421 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     6481 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     6541 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     6601 cagcgagtca gtgagcgagg aagcggaagg cgagagtagg gaactgccag gcatcaaact
     6661 aagcagaagg cccctgacgg atggcctttt tgcgtttcta caaactcttt ctgtgttgta
     6721 aaacgacggc cagtcttaag ctcgggcccc ctgggcggtt ctgataacga gtaatcgtta
     6781 atccgcaaat aacgtaaaaa cccgcttcgg cgggtttttt tatgggggga gtttagggaa
     6841 agagcatttg tcagaatatt taagggcgcc tgtcactttg cttgatatat gagaattatt
     6901 taaccttata aatgagaaaa aagcaacgca ctttaaataa gatacgttgc tttttcgatt
     6961 gatgaacacc tataattaaa ctattcatct attatttatg attttttgta tatacaatat
     7021 ttctagtttg ttaaagagaa ttaagaaaat aaatctcgaa aataataaag ggaaaatcag
     7081 tttttgatat caaaattata catgtcaacg ataatacaaa atataataca aactataaga
     7141 tgttatcagt atttattatc atttagaata aattttgtgt cgcccttaat tgtgagcgga
     7201 taacaattac gagcttcatg cacagtggcg ttgactcaat attggccatt agccatatta
     7261 ttcattggtt atatagcata aatcaatatt ggctattggc cattgcatac gttgtatcta
     7321 tatcataata tgtacattta tattggctca tgtccaatat gaccgccatg tt
//
