LOCUS       sequence-303133-        9016 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 9016)
  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 9016)
  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-CNTN1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..9016
                     /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             6358..6402
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6403..6420
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6560..6577)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6566..6677
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6795..6815)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6800..7129
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6980..7115
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7042..7061
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7177..7224
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7243..7842
                     /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(7243..7262)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7739..7759
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7972..8093
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8009..8028)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8063..8082
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8146..8937,8938..9006))
                     /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     8769..8788
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(9007..9016,1..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 cggccgccga gtttacctgg
     2761 tatagaagat atggccacgg cgtgtccgaa gaggacaaag gctttggccc catcttcgag
     2821 gaacagccca tcaacacaat ctaccccgag gaaagcctgg aaggcaaggt gtccctgaac
     2881 tgcagagcca gagcctctcc atttcctgtg tacaagtggc ggatgaacaa cggcgacgtg
     2941 gacctgacca gcgacagata tagcatggtc ggaggcaacc tggtcatcaa caaccccgac
     3001 aagcagaagg acgccggcat ctactactgc ctggccagca acaattacgg catggtccga
     3061 agcacagagg ccacactgag cttcggctat ctggacccat ttcctccaga ggaacggccc
     3121 gaagttcgcg tgaaagaagg caaaggcatg gtgctgctgt gcgaccctcc ttatcacttc
     3181 cccgacgacc tgagctacag atggctgctg aacgagttcc ccgtgttcat caccatggac
     3241 aagcggagat tcgtgtccca gaccaacggc aacctgtata tcgccaacgt ggaagccagc
     3301 gacaagggca actacagctg cttcgtgtct agccccagca tcaccaagag cgtgttcagc
     3361 aagttcatcc ctctgatccc cattcctgag cggaccacca agccttatcc tgccgatatc
     3421 gtggtgcagt tcaaggatgt gtacgccctg atgggccaga acgtgaccct ggaatgtttc
     3481 gccctgggca accctgtgcc agacattaga tggcggaagg tgctggaacc tatgcctagc
     3541 acagccgaga tctctacatc tggcgccgtg ctgaagatct ttaacatcca gctcgaggac
     3601 gagggcatct acgagtgcga ggccgagaac atcagaggca aggataagca ccaggccaga
     3661 atctacgtgc aggcctttcc tgagtgggtc gagcacatca acgacaccga agtggacatc
     3721 ggcagcgacc tgtattggcc ttgtgtggcc accggcaagc ccattcctac aatccggtgg
     3781 ctgaagaacg gctacgccta tcacaagggc gagctgagac tgtacgacgt gaccttcgag
     3841 aacgccggaa tgtaccagtg tatcgccgag aatacctacg gcgccatcta cgccaacgcc
     3901 gagctgaaaa ttctggccct ggctcctacc tttgagatga accccatgaa gaagaagatc
     3961 ctggccgcca aaggcggcag agtgatcatc gagtgcaagc ccaaagccgc tcctaagcct
     4021 aagttcagct ggtccaaggg caccgagtgg ctggtcaata gcagcagaat cctgatctgg
     4081 gaggacggca gcctggaaat caacaacatc accagaaacg acggcgggat ctacacctgt
     4141 ttcgccgaaa acaaccgcgg caaggccaac tctacaggca ccctcgtgat cacagacccc
     4201 accagaatca tcctggctcc aatcaacgcc gacatcaccg tgggcgagaa tgccacaatg
     4261 cagtgcgccg cctctttcga ccctgctctg gatctgacct tcgtgtggtc cttcaatggc
     4321 tacgtgatcg acttcaacaa agagaacatc cactaccagc ggaacttcat gctggacagc
     4381 aacggcgagc tgctgatcag aaatgcccag ctgaagcacg ccggacggta cacatgtaca
     4441 gcccagacca tcgtggataa cagcagcgcc tctgccgatc tggttgttag aggacctcct
     4501 ggaccacctg gcggcctgag aattgaggat atcagagcca caagcgtggc cctgacatgg
     4561 tccagaggca gcgataatca cagccccatc agcaagtaca ccatccagac caagaccatc
     4621 ctgagcgacg actggaagga tgccaagacc gatccaccta tcatcgaggg aaacatggaa
     4681 gccgccagag ccgtggatct gatcccttgg atggaatacg agttcagagt ggtggccact
     4741 aacacactcg gaagaggcga gcctagcatc cccagcaaca gaatcaaaac cgacggcgct
     4801 gcccctaatg tggccccttc tgatgttggc ggaggcggcg gaagaaacag agagctgaca
     4861 attacttggg cccctctgag cagagagtac cactacggca acaacttcgg ctacatcgtg
     4921 gccttcaagc ccttcgacgg cgaggaatgg aagaaagtga ccgtgaccaa tcctgacacc
     4981 ggcagatacg tgcacaagga cgagacaatg agcccctcca ccgccttcca agtgaaagtg
     5041 aaggccttta acaacaaagg cgacggccct tacagcctgg tggccgtgat caattctgcc
     5101 caggatgctc cttctgaggc ccctactgaa gtgggcgtga aagtgctgag cagcagcgag
     5161 atttctgtgc actgggaaca cgtgctggaa aagatcgtgg aaagctacca gatccggtac
     5221 tgggccgctc acgacaaaga agaggccgcc aatagagtgc aagtgacgag ccaagagtac
     5281 agcgccaggc tggaaaacct gctgcctgac acacagtact tcatcgaagt gggagcctgc
     5341 aatagcgccg gatgtggacc tcctagcgac atgatcgagg cctttaccaa gaaggcccct
     5401 cctagccagc ctcctagaat catcagctct gtcagaagcg gcagccggta catcatcacc
     5461 tgggatcatg tggtggccct gtccaatgag agcaccgtga caggctacaa ggtgctgtac
     5521 agacccgatg gccagcacga cggaaagctg tatagcaccc acaagcacag catcgaggtg
     5581 cccattccaa gagatggcga gtacgtggtg gaagtgcggg ctcattctga tggcggagat
     5641 ggggtcgtgt ctcaagttgg cgcgccgggc ggcgacaaga cccacacctg ccccccctgc
     5701 ccagctccag aactgctggg agggccttcc gtgttcctgt tcccccccaa gcccaaggac
     5761 accctgatga tcagcaggac ccccgaggtg acctgcgtgg tggtggccgt gtcccacgag
     5821 gacccagagg tgaagttcaa ctggtacgtg gacggcgtgg aggtgcacaa cgccaagacc
     5881 aagcccagag aggagcagta caacagcacc tacagggtgg tgtccgtgct gaccgtgctg
     5941 caccaggact ggctgaacgg caaagaatac aagtgcaaag tctccaacaa ggccctggct
     6001 gccccaatcg aaaagacaat cagcaaggcc aagggccagc cacgggagcc ccaggtgtac
     6061 accctgcccc ccagccggga ggagatgacc aagaaccagg tgtccctgac ctgtctggtg
     6121 aagggcttct accccagcga tatcgccgtg gagtgggaga gcaacggcca gcccgagaac
     6181 aactacaaga ccaccccccc agtgctggac agcgacggca gcttcttcct gtacagcaag
     6241 ctgaccgtgg acaagtccag gtggcagcag ggcaacgtgt tcagctgcag cgtgatgcac
     6301 gaggccctgc acaaccacta cacccagaag tccctgagcc tgagccccgg cggcagcggt
     6361 cttaatgata ttttcgaggc tcagaagatt gagtggcatg aacaccacca ccatcaccac
     6421 taaggccggc ctgtagtcac tggcacaacg cgttacccgg taagccaatc gggtatacac
     6481 ggtcgtcata ctgcagacag ggttcttcta ctttgcaaga tagtcttgag tagtaaaata
     6541 aatagataga gaaaatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     6601 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     6661 aatgaggaaa ttgcatcaca acactcaacc ctatctcggt ctattctttt gatttataag
     6721 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     6781 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     6841 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     6901 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     6961 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     7021 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     7081 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     7141 agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag
     7201 tatatcggca tagtataata cgacaaggtg aggaactaaa ccatgaccga gtacaagccc
     7261 acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg
     7321 ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc
     7381 accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc
     7441 gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg
     7501 gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag
     7561 caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc
     7621 accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc
     7681 ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacatc cgcgccccgc
     7741 aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa
     7801 ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacacgtgct acgagatttc
     7861 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     7921 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt
     7981 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     8041 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     8101 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcattacc aatgcttaat
     8161 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     8221 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagcg ctgcgatgat
     8281 accgcgagaa ccacgctcac cggctccgga tttatcagca ataaaccagc cagccggaag
     8341 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     8401 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccatcgc
     8461 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     8521 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     8581 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     8641 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     8701 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     8761 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     8821 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     8881 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     8941 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     9001 actcatattc ttcctt
//
