LOCUS       sequence-417859-        7060 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 7060)
  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 7060)
  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-NECTIN4-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..7060
                     /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             2636..2680
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2681..2698
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2838..2855)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2844..2955
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3073..3093)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3078..3407
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3258..3393
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3320..3339
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3455..3502
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3521..4120
                     /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(3521..3540)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4017..4037
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4250..4371
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4287..4306)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4341..4360
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4424..5215,5216..5284))
                     /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     5047..5066
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5285..5376)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5418..5437
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5443..5462
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5548..6136
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6037..6056
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6290..6307
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6405..6422
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6406..6422
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6878..6894
                     /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 ccggcgaact ggaaacatct gatgtggtta cagtggtgct
     1021 cggccaggat gccaagctgc cctgttttta cagaggcgac tctggcgagc aagtcggaca
     1081 agtggcttgg gctagagttg atgctggcga aggcgctcaa gaactggccc tgctgcactc
     1141 taagtacgga ctgcatgtgt ccccagccta cgagggaaga gttgaacagc cgcctcctcc
     1201 tagaaaccct ctggatggaa gcgtgctgct gagaaatgcc gtgcaggccg atgagggcga
     1261 gtacgagtgt agagtgtcta cctttccagc cggcagcttc caggccagac tgagactgag
     1321 agttctggtg cctcctctgc cttctctgaa ccctggacct gctctggaag aaggccaagg
     1381 actgacactg gccgcctctt gtacagccga aggatctcct gctcctagcg tgacctggga
     1441 taccgaagtg aagggcacca ccagcagcag aagcttcaag cacagtagat ctgccgccgt
     1501 gaccagcgag tttcatctgg tgcccagcag atccatgaac ggccagcctc tgacatgcgt
     1561 ggtgtctcat cctggactgc tgcaggacca gagaatcacc cacatcctgc acgtgtcctt
     1621 tctggccgag gcctctgtta gaggactgga agatcagaac ctgtggcaca tcggcagaga
     1681 aggggccatg ctgaagtgtc tgtctgaggg acagcctcct ccaagctaca actggacaag
     1741 actggatggc cctctgccta gcggagtcag agtggatggc gatacactgg gctttccacc
     1801 tctgacaaca gagcacagcg gcatctacgt gtgccacgtg tccaacgagt tcagcagcag
     1861 ggatagccaa gtgaccgtgg acgtgctgga ccctcaagag gattctggca aacaggtgga
     1921 cctggtgtcc gcctctggcg cgccgggcgg cgacaagacc cacacctgcc ccccctgccc
     1981 agctccagaa ctgctgggag ggccttccgt gttcctgttc ccccccaagc ccaaggacac
     2041 cctgatgatc agcaggaccc ccgaggtgac ctgcgtggtg gtggccgtgt cccacgagga
     2101 cccagaggtg aagttcaact ggtacgtgga cggcgtggag gtgcacaacg ccaagaccaa
     2161 gcccagagag gagcagtaca acagcaccta cagggtggtg tccgtgctga ccgtgctgca
     2221 ccaggactgg ctgaacggca aagaatacaa gtgcaaagtc tccaacaagg ccctggctgc
     2281 cccaatcgaa aagacaatca gcaaggccaa gggccagcca cgggagcccc aggtgtacac
     2341 cctgcccccc agccgggagg agatgaccaa gaaccaggtg tccctgacct gtctggtgaa
     2401 gggcttctac cccagcgata tcgccgtgga gtgggagagc aacggccagc ccgagaacaa
     2461 ctacaagacc acccccccag tgctggacag cgacggcagc ttcttcctgt acagcaagct
     2521 gaccgtggac aagtccaggt ggcagcaggg caacgtgttc agctgcagcg tgatgcacga
     2581 ggccctgcac aaccactaca cccagaagtc cctgagcctg agccccggcg gcagcggtct
     2641 taatgatatt ttcgaggctc agaagattga gtggcatgaa caccaccacc atcaccacta
     2701 aggccggcct gtagtcactg gcacaacgcg ttacccggta agccaatcgg gtatacacgg
     2761 tcgtcatact gcagacaggg ttcttctact ttgcaagata gtcttgagta gtaaaataaa
     2821 tagatagaga aaatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
     2881 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
     2941 tgaggaaatt gcatcacaac actcaaccct atctcggtct attcttttga tttataaggg
     3001 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     3061 aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     3121 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     3181 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     3241 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     3301 atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat
     3361 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag
     3421 cttgtatatc cattttcgga tctgatcagc acgtgttgac aattaatcat cggcatagta
     3481 tatcggcata gtataatacg acaaggtgag gaactaaacc atgaccgagt acaagcccac
     3541 ggtgcgcctc gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt
     3601 cgccgactac cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac
     3661 cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc
     3721 ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt
     3781 gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca
     3841 acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac
     3901 cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg
     3961 agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacatccg cgccccgcaa
     4021 cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg
     4081 accgcgcacc tggtgcatga cccgcaagcc cggtgcctga cacgtgctac gagatttcga
     4141 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     4201 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     4261 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     4321 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     4381 tataccgtcg acctctagct agagcttggc gtaatcatgg tcattaccaa tgcttaatca
     4441 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     4501 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagcgct gcgatgatac
     4561 cgcgagaacc acgctcaccg gctccggatt tatcagcaat aaaccagcca gccggaaggg
     4621 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     4681 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccatcgcta
     4741 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     4801 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     4861 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     4921 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     4981 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     5041 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     5101 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     5161 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     5221 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     5281 tcatattctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     5341 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggtcagtgtt acaaccaatt
     5401 aaccaattct gaacattatc gcgagcccat ttatacctga atatggctca taacacccct
     5461 tgctcatgac caaaatccct taacgtgagt tacgcgcgcg tcgttccact gagcgtcaga
     5521 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     5581 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     5641 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     5701 agtgtagccg tagttagccc accacttcaa gaactctgta gcaccgccta catacctcgc
     5761 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     5821 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     5881 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     5941 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     6001 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     6061 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     6121 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     6181 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     6241 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     6301 gagcgaggaa gcggaaggcg agagtaggga actgccaggc atcaaactaa gcagaaggcc
     6361 cctgacggat ggcctttttg cgtttctaca aactctttct gtgttgtaaa acgacggcca
     6421 gtcttaagct cgggccccct gggcggttct gataacgagt aatcgttaat ccgcaaataa
     6481 cgtaaaaacc cgcttcggcg ggttttttta tggggggagt ttagggaaag agcatttgtc
     6541 agaatattta agggcgcctg tcactttgct tgatatatga gaattattta accttataaa
     6601 tgagaaaaaa gcaacgcact ttaaataaga tacgttgctt tttcgattga tgaacaccta
     6661 taattaaact attcatctat tatttatgat tttttgtata tacaatattt ctagtttgtt
     6721 aaagagaatt aagaaaataa atctcgaaaa taataaaggg aaaatcagtt tttgatatca
     6781 aaattataca tgtcaacgat aatacaaaat ataatacaaa ctataagatg ttatcagtat
     6841 ttattatcat ttagaataaa ttttgtgtcg cccttaattg tgagcggata acaattacga
     6901 gcttcatgca cagtggcgtt gactcaatat tggccattag ccatattatt cattggttat
     6961 atagcataaa tcaatattgg ctattggcca ttgcatacgt tgtatctata tcataatatg
     7021 tacatttata ttggctcatg tccaatatga ccgccatgtt
//
