LOCUS       sequence-420993-        7003 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 7003)
  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 7003)
  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-MADCAM1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..7003
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        165..544
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        545..748
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     699..719
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     745..769
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        979..997
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     979..996
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     1010..1029
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2743..2787
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2788..2805
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2945..2962)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2951..3062
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3180..3200)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3185..3514
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3365..3500
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3427..3446
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3562..3609
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3628..4227
                     /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(3628..3647)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4124..4144
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4357..4478
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4394..4413)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4448..4467
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4531..5322,5323..5391))
                     /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     5154..5173
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5392..5483)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5525..5544
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5550..5569
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5655..6243
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6144..6163
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6397..6414
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6512..6529
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6513..6529
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6985..7001
                     /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 acgagcttca tgcacagtgg cgttgactca atattggcca ttagccatat tattcattgg
       61 ttatatagca taaatcaata ttggctattg gccattgcat acgttgtatc tatatcataa
      121 tatgtacatt tatattggct catgtccaat atgaccgcca tgttggcatt gattattgac
      181 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
      241 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      301 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      361 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      421 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      481 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      541 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      601 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      661 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      721 acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagatcg cctggagacg
      781 ccatccacgc tgttttgacc tccatagaag acaccgggac cgatccagcc tccgcggccg
      841 ggaacggtgc attggaacgc ggattccccg tgccaagagt gacgtaagta ccgcctatag
      901 agtctatagg cccaccccct tggcttcgtt agaacgcggc tacaattaat acataacctt
      961 atgtatcata cacatacgat ttaggtgaca ctatagaata acatccactt tgcctttctc
     1021 tccacaggtg tccactccca ggtccaactg cacctcggtt ctatcgaaaa cgcgcctgct
     1081 agcgccacca tgaagaccat cattgccctg agctacatct tctgcctggt gttcgctgcg
     1141 gccgcccaat ctctgcaagt gaagcctctt caggtggaac ctcctgagcc tgtggttgct
     1201 gttgctctgg gagcttccag acagctgacc tgtagactgg cctgtgccga tagaggcgct
     1261 tctgttcaat ggcgcggact ggatacaagc ctgggagccg ttcagagcga taccggcaga
     1321 tctgtgctga cagtgcggaa tgcctctctg tctgccgccg gaacaagagt gtgtgtggga
     1381 tcttgtggcg gcagaacctt tcagcacaca gtgcagctgc tggtgtacgc ctttcctgac
     1441 cagctgacag tttctcctgc tgctctggtg cctggcgatc ctgaagtggc ctgtactgcc
     1501 cacaaagtga cccctgtgga ccccaatgct ctgagctttt ctctgctcgt cggcggccaa
     1561 gaactggaag gtgctcaagc tctgggcccc gaggttcaag aggaagaaga agaacctcag
     1621 ggcgacgagg acgtgctgtt cagagttaca gaaagatggc ggctgcctcc tctgggaaca
     1681 cctgttcctc cagctctgta ctgtcaggcc acaatgagac tgcctggcct ggaactgagc
     1741 cacagacagg ctattccagt gctgcacagc cctacaagcc ctgagcctcc tgatacaaca
     1801 agcccagaga gccccgatac cacctctcct gagagccctg acaccacaag ccaagagcct
     1861 ccagacacta catccccaga gccacctgac aagacctctc cagaacctgc tcctcagcag
     1921 ggcagcacac acacacctag aagccctggc agcaccagaa ccagaaggcc tgaaattagc
     1981 caggctggcc ctacacaggg cgaagtgatc ccaacaggat ctagcaagcc tgccggcgac
     2041 caaggcgcgc cgggcggcga caagacccac acctgccccc cctgcccagc tccagaactg
     2101 ctgggagggc cttccgtgtt cctgttcccc cccaagccca aggacaccct gatgatcagc
     2161 aggacccccg aggtgacctg cgtggtggtg gccgtgtccc acgaggaccc agaggtgaag
     2221 ttcaactggt acgtggacgg cgtggaggtg cacaacgcca agaccaagcc cagagaggag
     2281 cagtacaaca gcacctacag ggtggtgtcc gtgctgaccg tgctgcacca ggactggctg
     2341 aacggcaaag aatacaagtg caaagtctcc aacaaggccc tggctgcccc aatcgaaaag
     2401 acaatcagca aggccaaggg ccagccacgg gagccccagg tgtacaccct gccccccagc
     2461 cgggaggaga tgaccaagaa ccaggtgtcc ctgacctgtc tggtgaaggg cttctacccc
     2521 agcgatatcg ccgtggagtg ggagagcaac ggccagcccg agaacaacta caagaccacc
     2581 cccccagtgc tggacagcga cggcagcttc ttcctgtaca gcaagctgac cgtggacaag
     2641 tccaggtggc agcagggcaa cgtgttcagc tgcagcgtga tgcacgaggc cctgcacaac
     2701 cactacaccc agaagtccct gagcctgagc cccggcggca gcggtcttaa tgatattttc
     2761 gaggctcaga agattgagtg gcatgaacac caccaccatc accactaagg ccggcctgta
     2821 gtcactggca caacgcgtta cccggtaagc caatcgggta tacacggtcg tcatactgca
     2881 gacagggttc ttctactttg caagatagtc ttgagtagta aaataaatag atagagaaaa
     2941 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     3001 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     3061 tcacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     3121 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg
     3181 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3241 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     3301 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     3361 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     3421 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     3481 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     3541 tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat cggcatagta
     3601 taatacgaca aggtgaggaa ctaaaccatg accgagtaca agcccacggt gcgcctcgcc
     3661 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc
     3721 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa
     3781 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc
     3841 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc
     3901 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc
     3961 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg
     4021 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc
     4081 gagcgcgccg gggtgcccgc cttcctggag acatccgcgc cccgcaacct ccccttctac
     4141 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg
     4201 tgcatgaccc gcaagcccgg tgcctgacac gtgctacgag atttcgattc caccgccgcc
     4261 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     4321 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
     4381 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     4441 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
     4501 tctagctaga gcttggcgta atcatggtca ttaccaatgc ttaatcagtg aggcacctat
     4561 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     4621 tacgatacgg gagggcttac catctggccc cagcgctgcg atgataccgc gagaaccacg
     4681 ctcaccggct ccggatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     4741 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     4801 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc atcgctacag gcatcgtggt
     4861 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     4921 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     4981 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     5041 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     5101 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     5161 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     5221 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     5281 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     5341 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tattcttcct
     5401 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     5461 atgtatttag aaaaataaac aaataggggt cagtgttaca accaattaac caattctgaa
     5521 cattatcgcg agcccattta tacctgaata tggctcataa caccccttgc tcatgaccaa
     5581 aatcccttaa cgtgagttac gcgcgcgtcg ttccactgag cgtcagaccc cgtagaaaag
     5641 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     5701 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     5761 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     5821 ttagcccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     5881 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     5941 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     6001 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     6061 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     6121 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     6181 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     6241 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     6301 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     6361 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     6421 gaaggcgaga gtagggaact gccaggcatc aaactaagca gaaggcccct gacggatggc
     6481 ctttttgcgt ttctacaaac tctttctgtg ttgtaaaacg acggccagtc ttaagctcgg
     6541 gccccctggg cggttctgat aacgagtaat cgttaatccg caaataacgt aaaaacccgc
     6601 ttcggcgggt ttttttatgg ggggagttta gggaaagagc atttgtcaga atatttaagg
     6661 gcgcctgtca ctttgcttga tatatgagaa ttatttaacc ttataaatga gaaaaaagca
     6721 acgcacttta aataagatac gttgcttttt cgattgatga acacctataa ttaaactatt
     6781 catctattat ttatgatttt ttgtatatac aatatttcta gtttgttaaa gagaattaag
     6841 aaaataaatc tcgaaaataa taaagggaaa atcagttttt gatatcaaaa ttatacatgt
     6901 caacgataat acaaaatata atacaaacta taagatgtta tcagtattta ttatcattta
     6961 gaataaattt tgtgtcgccc ttaattgtga gcggataaca att
//
