LOCUS       sequence-401535-        6976 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 6976)
  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 6976)
  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-TIMD4-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6976
                     /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             2552..2596
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2597..2614
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2754..2771)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2760..2871
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2989..3009)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2994..3323
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3174..3309
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3236..3255
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3371..3418
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3437..4036
                     /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(3437..3456)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3933..3953
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4166..4287
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4203..4222)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4257..4276
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4340..5131,5132..5200))
                     /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     4963..4982
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5201..5292)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5334..5353
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5359..5378
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5464..6052
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5953..5972
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6206..6223
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6321..6338
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6322..6338
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6794..6810
                     /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 ccgaaacagt ggttacagaa gtgctgggcc acagagtgac
     1021 cctgccttgt ctgtacagca gctggtccca caacagcaac agcatgtgct ggggcaaaga
     1081 tcagtgcccc tacagcggat gcaaagaggc cctgatcaga accgacggca tgagagtgac
     1141 aagcagaaag agcgccaagt accggctgca gggaacaatc cctagaggcg acgtgtccct
     1201 gaccattctg aaccctagcg agagcgatag cggcgtgtac tgctgtagaa tcgaggtgcc
     1261 cggctggttc aacgacgtga agatcaacgt gcggctgaac ctgcagagag ccagcaccac
     1321 aacacacaga accgccacaa ccaccaccag acggaccaca acaacaagcc ccaccaccac
     1381 aagacagatg accactactc ctgccgctct gcctaccacc gtggtcacaa cacctgatct
     1441 gaccacaggc acccctctgc agatgacaac aatcgccgtg ttcaccaccg ccaatacctg
     1501 cctgagcctg acacctagca cactgcctga agaagccaca ggcctgctga cccctgagcc
     1561 ttctaaagag ggacctatcc tgaccgccga gagcgagaca gtgctgcctt ctgatagctg
     1621 gtccagcgtg gaaagcacca gcgctgatac cgtgctgctg accagcaaag aaagcaaagt
     1681 gtgggacctg cctagcacca gccacgtgtc catgtggaaa accagcgact ccgtgtcatc
     1741 tcctcagcca ggcgcttctg atacagccgt gcctgagcag aacaagacca ccaagacagg
     1801 ccagatggac ggcatcccca tgagcatgaa gaacgagatg cccatcagcc aaggcgcgcc
     1861 gggcggcgac aagacccaca cctgcccccc ctgcccagct ccagaactgc tgggagggcc
     1921 ttccgtgttc ctgttccccc ccaagcccaa ggacaccctg atgatcagca ggacccccga
     1981 ggtgacctgc gtggtggtgg ccgtgtccca cgaggaccca gaggtgaagt tcaactggta
     2041 cgtggacggc gtggaggtgc acaacgccaa gaccaagccc agagaggagc agtacaacag
     2101 cacctacagg gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaaaga
     2161 atacaagtgc aaagtctcca acaaggccct ggctgcccca atcgaaaaga caatcagcaa
     2221 ggccaagggc cagccacggg agccccaggt gtacaccctg ccccccagcc gggaggagat
     2281 gaccaagaac caggtgtccc tgacctgtct ggtgaagggc ttctacccca gcgatatcgc
     2341 cgtggagtgg gagagcaacg gccagcccga gaacaactac aagaccaccc ccccagtgct
     2401 ggacagcgac ggcagcttct tcctgtacag caagctgacc gtggacaagt ccaggtggca
     2461 gcagggcaac gtgttcagct gcagcgtgat gcacgaggcc ctgcacaacc actacaccca
     2521 gaagtccctg agcctgagcc ccggcggcag cggtcttaat gatattttcg aggctcagaa
     2581 gattgagtgg catgaacacc accaccatca ccactaaggc cggcctgtag tcactggcac
     2641 aacgcgttac ccggtaagcc aatcgggtat acacggtcgt catactgcag acagggttct
     2701 tctactttgc aagatagtct tgagtagtaa aataaataga tagagaaaat cagcctcgac
     2761 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     2821 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cacaacactc
     2881 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     2941 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc
     3001 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3061 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3121 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     3181 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     3241 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt
     3301 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg
     3361 atcagcacgt gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa
     3421 ggtgaggaac taaaccatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga
     3481 cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca
     3541 caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac
     3601 gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt
     3661 ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat
     3721 ggccgagttg agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc
     3781 gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca
     3841 gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg
     3901 ggtgcccgcc ttcctggaga catccgcgcc ccgcaacctc cccttctacg agcggctcgg
     3961 cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg
     4021 caagcccggt gcctgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag
     4081 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     4141 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     4201 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     4261 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     4321 cttggcgtaa tcatggtcat taccaatgct taatcagtga ggcacctatc tcagcgatct
     4381 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     4441 agggcttacc atctggcccc agcgctgcga tgataccgcg agaaccacgc tcaccggctc
     4501 cggatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     4561 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     4621 cagttaatag tttgcgcaac gttgttgcca tcgctacagg catcgtggtg tcacgctcgt
     4681 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     4741 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     4801 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     4861 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     4921 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     4981 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     5041 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     5101 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     5161 aaaagggaat aagggcgaca cggaaatgtt gaatactcat attcttcctt tttcaatatt
     5221 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     5281 aaaataaaca aataggggtc agtgttacaa ccaattaacc aattctgaac attatcgcga
     5341 gcccatttat acctgaatat ggctcataac accccttgct catgaccaaa atcccttaac
     5401 gtgagttacg cgcgcgtcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     5461 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     5521 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     5581 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt tagcccacca
     5641 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     5701 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     5761 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     5821 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     5881 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     5941 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     6001 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     6061 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     6121 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     6181 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aaggcgagag
     6241 tagggaactg ccaggcatca aactaagcag aaggcccctg acggatggcc tttttgcgtt
     6301 tctacaaact ctttctgtgt tgtaaaacga cggccagtct taagctcggg ccccctgggc
     6361 ggttctgata acgagtaatc gttaatccgc aaataacgta aaaacccgct tcggcgggtt
     6421 tttttatggg gggagtttag ggaaagagca tttgtcagaa tatttaaggg cgcctgtcac
     6481 tttgcttgat atatgagaat tatttaacct tataaatgag aaaaaagcaa cgcactttaa
     6541 ataagatacg ttgctttttc gattgatgaa cacctataat taaactattc atctattatt
     6601 tatgattttt tgtatataca atatttctag tttgttaaag agaattaaga aaataaatct
     6661 cgaaaataat aaagggaaaa tcagtttttg atatcaaaat tatacatgtc aacgataata
     6721 caaaatataa tacaaactat aagatgttat cagtatttat tatcatttag aataaatttt
     6781 gtgtcgccct taattgtgag cggataacaa ttacgagctt catgcacagt ggcgttgact
     6841 caatattggc cattagccat attattcatt ggttatatag cataaatcaa tattggctat
     6901 tggccattgc atacgttgta tctatatcat aatatgtaca tttatattgg ctcatgtcca
     6961 atatgaccgc catgtt
//
