LOCUS       sequence-407026-        7540 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 7540)
  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 7540)
  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-LRFN4-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..7540
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     124..143
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     149..168
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      254..842
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     743..762
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     996..1013
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1111..1128
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1112..1128
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1584..1600
                     /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)."
     enhancer        1767..2146
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2147..2350
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2301..2321
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2347..2371
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2581..2599
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2581..2598
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2612..2631
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             4954..4998
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4999..5016
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5156..5173)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5162..5273
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5391..5411)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5435..5645
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        5457..5653
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5504..5639
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5566..5585
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5701..5748
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5767..6366
                     /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(5767..5786)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6263..6283
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6496..6617
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6533..6552)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6587..6606
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6670..7461,7462..7530))
                     /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     7293..7312
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7531..7540,1..82))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
       61 tgtatttaga aaaataaaca aataggggtc agtgttacaa ccaattaacc aattctgaac
      121 attatcgcga gcccatttat acctgaatat ggctcataac accccttgct catgaccaaa
      181 atcccttaac gtgagttacg cgcgcgtcgt tccactgagc gtcagacccc gtagaaaaga
      241 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
      301 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
      361 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
      421 tagcccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
      481 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
      541 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
      601 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
      661 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
      721 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
      781 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
      841 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
      901 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
      961 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     1021 aaggcgagag tagggaactg ccaggcatca aactaagcag aaggcccctg acggatggcc
     1081 tttttgcgtt tctacaaact ctttctgtgt tgtaaaacga cggccagtct taagctcggg
     1141 ccccctgggc ggttctgata acgagtaatc gttaatccgc aaataacgta aaaacccgct
     1201 tcggcgggtt tttttatggg gggagtttag ggaaagagca tttgtcagaa tatttaaggg
     1261 cgcctgtcac tttgcttgat atatgagaat tatttaacct tataaatgag aaaaaagcaa
     1321 cgcactttaa ataagatacg ttgctttttc gattgatgaa cacctataat taaactattc
     1381 atctattatt tatgattttt tgtatataca atatttctag tttgttaaag agaattaaga
     1441 aaataaatct cgaaaataat aaagggaaaa tcagtttttg atatcaaaat tatacatgtc
     1501 aacgataata caaaatataa tacaaactat aagatgttat cagtatttat tatcatttag
     1561 aataaatttt gtgtcgccct taattgtgag cggataacaa ttacgagctt catgcacagt
     1621 ggcgttgact caatattggc cattagccat attattcatt ggttatatag cataaatcaa
     1681 tattggctat tggccattgc atacgttgta tctatatcat aatatgtaca tttatattgg
     1741 ctcatgtcca atatgaccgc catgttggca ttgattattg actagttatt aatagtaatc
     1801 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     1861 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     1921 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     1981 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     2041 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     2101 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     2161 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     2221 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     2281 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     2341 aagcagagct cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga
     2401 cctccataga agacaccggg accgatccag cctccgcggc cgggaacggt gcattggaac
     2461 gcggattccc cgtgccaaga gtgacgtaag taccgcctat agagtctata ggcccacccc
     2521 cttggcttcg ttagaacgcg gctacaatta atacataacc ttatgtatca tacacatacg
     2581 atttaggtga cactatagaa taacatccac tttgcctttc tctccacagg tgtccactcc
     2641 caggtccaac tgcacctcgg ttctatcgaa aacgcgcctg ctagcgccac catgaagacc
     2701 atcattgccc tgagctacat cttctgcctg gtgttcgctg cggccgcctg tcctttgcct
     2761 tgcgtgtgtc agaatctgag cgagagcctg agcaccctgt gtgcccatag aggcctgctg
     2821 ttcgtgcctc ctaatgtgga cagacggacc gtggaactga gactggccga caacttcatc
     2881 caggctctgg gccctccaga cttcagaaac atgaccggcc tggtggatct gaccctgagc
     2941 agaaacgcca tcaccagaat tggcgccaga gccttcggcg acctggaaag cctgagaagc
     3001 ctgcacctgg acggcaacag actggtggaa ctcggcaccg gatctctgag aggccctgtg
     3061 aatctgcagc acctgatcct gagcggcaac cagctgggaa gaattgcccc tggcgccttc
     3121 gacgacttcc tggaatctct ggaagatctg gacctgagct acaacaacct gagacaggtg
     3181 ccatgggccg gcattggagc tatgcctgct ctgcacaccc tgaacctgga ccacaacctg
     3241 atcgatgctc tgcctcctgg cgcttttgct cagctgggac aactgagcag actggacctc
     3301 accagcaaca ggctggctac tctggctccc gatcctctgt ttagcagagg cagagatgcc
     3361 gaggcctctc ctgctcctct ggtgctgagc tttagcggca atcccctgca ctgcaactgc
     3421 gagctgctgt ggcttcggag actggctaga cccgacgatc tggaaacctg cgcttctcct
     3481 cctggactgg ccggcagata cttttgggct gtgcctgagg gcgagttcag ctgcgaacct
     3541 cctctgatcg ccagacacac ccagagactg tgggtgctcg aaggacagag ggccacactg
     3601 agatgtagag ccctgggaga tcccgctcct accatgcatt gggtcggacc cgatgataga
     3661 ctcgtgggca acagcagcag agccagggcc tttcctaatg gcaccctgga aatcggcgtt
     3721 accggtgctg gcgacgctgg cggatacacc tgtatcgcta caaatcctgc cggcgaggcc
     3781 accgctagag ttgaactgag agttctggcc ctgcctcacg gcggaaatag ctctgctgaa
     3841 ggtggcagac ccggaccttc tgatattgcc gcctctgcca gaacagccgc tgaaggcgag
     3901 ggaacactgg aatctgaacc cgccgtgcaa gtgaccgaag tgacagccac atctggcctg
     3961 gtgtcttggg gacctggtag acctgctgat cccgtgtgga tgttccagat ccagtacaac
     4021 agctccgagg acgagacact gatctacaga atcgtgcccg ccagcagcca ccactttctg
     4081 ctgaaacatc tggtgcctgg cgccgactac gatctgtgtc tgctggctct ttctcctgcc
     4141 gccggtccta gtgatctgac cgctacaaga ctgctgggct gcgcccactt tagcaccttg
     4201 cctgcttctc cactgtgtca cgccctgcag gctcatgtgc tcggaggaac acttggcgcg
     4261 ccgggcggcg acaagaccca cacctgcccc ccctgcccag ctccagaact gctgggaggg
     4321 ccttccgtgt tcctgttccc ccccaagccc aaggacaccc tgatgatcag caggaccccc
     4381 gaggtgacct gcgtggtggt ggccgtgtcc cacgaggacc cagaggtgaa gttcaactgg
     4441 tacgtggacg gcgtggaggt gcacaacgcc aagaccaagc ccagagagga gcagtacaac
     4501 agcacctaca gggtggtgtc cgtgctgacc gtgctgcacc aggactggct gaacggcaaa
     4561 gaatacaagt gcaaagtctc caacaaggcc ctggctgccc caatcgaaaa gacaatcagc
     4621 aaggccaagg gccagccacg ggagccccag gtgtacaccc tgccccccag ccgggaggag
     4681 atgaccaaga accaggtgtc cctgacctgt ctggtgaagg gcttctaccc cagcgatatc
     4741 gccgtggagt gggagagcaa cggccagccc gagaacaact acaagaccac ccccccagtg
     4801 ctggacagcg acggcagctt cttcctgtac agcaagctga ccgtggacaa gtccaggtgg
     4861 cagcagggca acgtgttcag ctgcagcgtg atgcacgagg ccctgcacaa ccactacacc
     4921 cagaagtccc tgagcctgag ccccggcggc agcggtctta atgatatttt cgaggctcag
     4981 aagattgagt ggcatgaaca ccaccaccat caccactaag gccggcctgt agtcactggc
     5041 acaacgcgtt acccggtaag ccaatcgggt atacacggtc gtcatactgc agacagggtt
     5101 cttctacttt gcaagatagt cttgagtagt aaaataaata gatagagaaa atcagcctcg
     5161 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     5221 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcacaacac
     5281 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     5341 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg
     5401 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     5461 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     5521 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     5581 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     5641 aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcagcacgt
     5701 gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa ggtgaggaac
     5761 taaaccatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg
     5821 gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat
     5881 ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg
     5941 ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg
     6001 ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg
     6061 agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc
     6121 aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt
     6181 ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc
     6241 ttcctggaga catccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc
     6301 accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt
     6361 gcctgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     6421 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     6481 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     6541 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     6601 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     6661 tcatggtcat taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6721 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6781 atctggcccc agcgctgcga tgataccgcg agaaccacgc tcaccggctc cggatttatc
     6841 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6901 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6961 tttgcgcaac gttgttgcca tcgctacagg catcgtggtg tcacgctcgt cgtttggtat
     7021 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     7081 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     7141 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     7201 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     7261 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     7321 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     7381 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     7441 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7501 aagggcgaca cggaaatgtt gaatactcat attcttcctt
//
