LOCUS       sequence-412906-        8203 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 8203)
  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 8203)
  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-NCAM1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8203
                     /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             5545..5589
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5590..5607
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5747..5764)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5753..5864
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5982..6002)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5987..6316
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6167..6302
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6229..6248
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6364..6411
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6430..7029
                     /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(6430..6449)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6926..6946
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7159..7280
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7196..7215)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7250..7269
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7333..8124,8125..8193))
                     /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     7956..7975
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8194..8203,1..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 cggccgcctt gcaagtggat
     2761 attgtgccta gccagggcga gatcagcgtg ggcgagtcta agttcttcct gtgccaagtg
     2821 gccggcgacg ccaaggacaa ggatatcagc tggttcagcc ccaacggcga gaagctgacc
     2881 cctaaccagc agagaatcag cgtcgtgtgg aacgacgaca gcagcagcac cctgaccatc
     2941 tacaacgcca acatcgacga cgccggcatc tataagtgtg tggtcacagg cgaggacggc
     3001 agcgagtctg aagccaccgt gaacgtgaag atctttcaga aactgatgtt caagaacgcc
     3061 ccgacgcctc aagagttcag agaaggcgag gatgccgtga tcgtgtgcga cgttgtcagc
     3121 agcctgcctc ctaccatcat ctggaagcac aagggccgcg acgtgatcct gaagaaagac
     3181 gtgcggttca tcgtgctgag caacaactac ctgcagatcc ggggcatcaa gaaaaccgac
     3241 gagggcacct acagatgcga aggcagaatc ctggccagag gggagatcaa cttcaaggac
     3301 atccaagtga tcgtgaacgt cccacctacc atccaggcca gacagaacat cgtgaatgcc
     3361 accgccaatc tgggccagag cgtgaccctt gtgtgtgatg ccgagggctt tcccgagcct
     3421 accatgagct ggacaaagga cggcgagcag atcgagcaag aagaggacga cgagaagtac
     3481 atcttcagcg acgactccag ccagctgacc atcaagaagg tggacaagaa cgacgaggcc
     3541 gagtacatct gtatcgccga gaacaaggcc ggcgaacagg atgccacaat ccacctgaag
     3601 gtgttcgcca agcctaagat cacctacgtg gaaaaccaga ccgccatgga actggaagaa
     3661 caagtgaccc tgacctgcga ggccagcggc gatcctattc cttctatcac ctggcggacc
     3721 agcaccagaa acatcagcag cgaggaaaag gccagctgga cccggcctga gaagcaagag
     3781 acactggatg gccacatggt cgtgcggagc catgccagag ttagcagcct gacactgaag
     3841 tccatccagt acacagacgc cggcgagtat atctgcaccg ccagcaatac catcggccag
     3901 gacagccaga gcatgtacct ggaagtgcag tacgccccta agctgcaggg acctgtggcc
     3961 gtgtatacct gggagggcaa tcaagtgaac atcacatgcg aagtgttcgc ttaccccagc
     4021 gccaccatta gctggtttag agatggccag ctgctgccca gcagcaacta cagcaacatc
     4081 aagatctaca acacccctag cgccagctat ctggaagtga ccccagacag cgagaacgac
     4141 ttcggcaact acaattgcac cgccgtgaac cggatcggcc aagagagcct ggaattcatc
     4201 ctggtgcagg ccgacacacc tagcagccct tctatcgacc aggtggaacc ctacagcagc
     4261 acagcccagg tgcagttcga tgagcctgaa gctacaggcg gcgtgcccat cctgaagtat
     4321 aaggccgagt ggcgcgctgt gggcgaagag gtttggcaca gcaagtggta cgacgccaaa
     4381 gaggccagca tggaaggcat cgtgaccatc gtgggcctga agcctgagac aacatatgcc
     4441 gtcagactgg ccgctctgaa cggcaaagga ctgggcgaaa ttagcgccgc cagcgagttt
     4501 aagacccagc ctgttcaggg cgagcccagc gctcctaaac tggaaggaca gatgggagaa
     4561 gatggcaaca gcatcaaagt gaacctgatc aagcaggacg acggcggcag ccccatcaga
     4621 cactatctcg tcagatacag agccctgagc agcgagtgga agcccgagat cagactgcct
     4681 agcggctctg accacgtgat gctgaaaagc ctggactgga acgccgagta cgaggtgtac
     4741 gtggtggccg aaaatcagca gggcaagagc aaggccgctc acttcgtgtt cagaacaagc
     4801 gcccagccta ccgccattcc tgccaatgga tctccaacca gcggaggcgc gccgggcggc
     4861 gacaagaccc acacctgccc cccctgccca gctccagaac tgctgggagg gccttccgtg
     4921 ttcctgttcc cccccaagcc caaggacacc ctgatgatca gcaggacccc cgaggtgacc
     4981 tgcgtggtgg tggccgtgtc ccacgaggac ccagaggtga agttcaactg gtacgtggac
     5041 ggcgtggagg tgcacaacgc caagaccaag cccagagagg agcagtacaa cagcacctac
     5101 agggtggtgt ccgtgctgac cgtgctgcac caggactggc tgaacggcaa agaatacaag
     5161 tgcaaagtct ccaacaaggc cctggctgcc ccaatcgaaa agacaatcag caaggccaag
     5221 ggccagccac gggagcccca ggtgtacacc ctgcccccca gccgggagga gatgaccaag
     5281 aaccaggtgt ccctgacctg tctggtgaag ggcttctacc ccagcgatat cgccgtggag
     5341 tgggagagca acggccagcc cgagaacaac tacaagacca cccccccagt gctggacagc
     5401 gacggcagct tcttcctgta cagcaagctg accgtggaca agtccaggtg gcagcagggc
     5461 aacgtgttca gctgcagcgt gatgcacgag gccctgcaca accactacac ccagaagtcc
     5521 ctgagcctga gccccggcgg cagcggtctt aatgatattt tcgaggctca gaagattgag
     5581 tggcatgaac accaccacca tcaccactaa ggccggcctg tagtcactgg cacaacgcgt
     5641 tacccggtaa gccaatcggg tatacacggt cgtcatactg cagacagggt tcttctactt
     5701 tgcaagatag tcttgagtag taaaataaat agatagagaa aatcagcctc gactgtgcct
     5761 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     5821 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcacaaca ctcaacccta
     5881 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     5941 atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg
     6001 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     6061 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     6121 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     6181 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     6241 ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     6301 cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcagca
     6361 cgtgttgaca attaatcatc ggcatagtat atcggcatag tataatacga caaggtgagg
     6421 aactaaacca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
     6481 agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
     6541 gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
     6601 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc
     6661 acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag
     6721 ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg
     6781 cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag
     6841 ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc
     6901 gccttcctgg agacatccgc gccccgcaac ctccccttct acgagcggct cggcttcacc
     6961 gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc
     7021 ggtgcctgac acgtgctacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     7081 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     7141 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     7201 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     7261 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     7321 taatcatggt cattaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     7381 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     7441 accatctggc cccagcgctg cgatgatacc gcgagaacca cgctcaccgg ctccggattt
     7501 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     7561 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     7621 tagtttgcgc aacgttgttg ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     7681 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     7741 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     7801 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     7861 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     7921 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     7981 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     8041 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     8101 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     8161 aataagggcg acacggaaat gttgaatact catattcttc ctt
//
