LOCUS       sequence-493105-        7531 bp    DNA     circular SYN 18-NOV-2025
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to COMP5AP-AviTag-9xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7531)
  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 7531)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 18, 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-CTLA4-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7531
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(87..104)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    93..204
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(322..342)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        327..656
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      507..642
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     569..588
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        704..751
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             770..1369
                     /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(770..789)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1266..1286
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1499..1620
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1536..1555)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1590..1609
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(1673..2464,2465..2533))
                     /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     2296..2315
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2534..2625)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2667..2686
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2692..2711
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2797..3385
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3286..3305
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3539..3556
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     3654..3671
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3655..3671
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4127..4143
                     /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        4310..4689
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4690..4893
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4844..4864
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4890..4914
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        5124..5142
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     5124..5141
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     5155..5174
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             5724..5747
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             7407..7451
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             7452..7478
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
ORIGIN
        1 tatacacggt cgtcatactg cagacagggt tcttctactt tgcaagatag tcttgagtag
       61 taaaataaat agatagagaa aatcagcctc gactgtgcct tctagttgcc agccatctgt
      121 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
      181 ctaataaaat gaggaaattg catcacaaca ctcaacccta tctcggtcta ttcttttgat
      241 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
      301 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
      361 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
      421 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
      481 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
      541 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
      601 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
      661 tcccgggagc ttgtatatcc attttcggat ctgatcagca cgtgttgaca attaatcatc
      721 ggcatagtat atcggcatag tataatacga caaggtgagg aactaaacca tgaccgagta
      781 caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc
      841 cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga
      901 gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt
      961 gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc
     1021 gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc
     1081 cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt
     1141 cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt
     1201 gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacatccgc
     1261 gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt
     1321 gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac acgtgctacg
     1381 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     1441 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     1501 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     1561 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     1621 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt cattaccaat
     1681 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     1741 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagcgctg
     1801 cgatgatacc gcgagaacca cgctcaccgg ctccggattt atcagcaata aaccagccag
     1861 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     1921 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     1981 ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     2041 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     2101 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     2161 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     2221 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     2281 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     2341 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     2401 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     2461 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     2521 gttgaatact catattcttc ctttttcaat attattgaag catttatcag ggttattgtc
     2581 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gtcagtgtta
     2641 caaccaatta accaattctg aacattatcg cgagcccatt tatacctgaa tatggctcat
     2701 aacacccctt gctcatgacc aaaatccctt aacgtgagtt acgcgcgcgt cgttccactg
     2761 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     2821 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     2881 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     2941 tgttcttcta gtgtagccgt agttagccca ccacttcaag aactctgtag caccgcctac
     3001 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     3061 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     3121 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     3181 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     3241 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     3301 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     3361 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     3421 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     3481 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     3541 cgagtcagtg agcgaggaag cggaaggcga gagtagggaa ctgccaggca tcaaactaag
     3601 cagaaggccc ctgacggatg gcctttttgc gtttctacaa actctttctg tgttgtaaaa
     3661 cgacggccag tcttaagctc gggccccctg ggcggttctg ataacgagta atcgttaatc
     3721 cgcaaataac gtaaaaaccc gcttcggcgg gtttttttat ggggggagtt tagggaaaga
     3781 gcatttgtca gaatatttaa gggcgcctgt cactttgctt gatatatgag aattatttaa
     3841 ccttataaat gagaaaaaag caacgcactt taaataagat acgttgcttt ttcgattgat
     3901 gaacacctat aattaaacta ttcatctatt atttatgatt ttttgtatat acaatatttc
     3961 tagtttgtta aagagaatta agaaaataaa tctcgaaaat aataaaggga aaatcagttt
     4021 ttgatatcaa aattatacat gtcaacgata atacaaaata taatacaaac tataagatgt
     4081 tatcagtatt tattatcatt tagaataaat tttgtgtcgc ccttaattgt gagcggataa
     4141 caattacgag cttcatgcac agtggcgttg actcaatatt ggccattagc catattattc
     4201 attggttata tagcataaat caatattggc tattggccat tgcatacgtt gtatctatat
     4261 cataatatgt acatttatat tggctcatgt ccaatatgac cgccatgttg gcattgatta
     4321 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
     4381 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
     4441 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
     4501 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
     4561 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
     4621 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
     4681 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
     4741 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
     4801 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
     4861 cgtgtacggt gggaggtcta tataagcaga gctcgtttag tgaaccgtca gatcgcctgg
     4921 agacgccatc cacgctgttt tgacctccat agaagacacc gggaccgatc cagcctccgc
     4981 ggccgggaac ggtgcattgg aacgcggatt ccccgtgcca agagtgacgt aagtaccgcc
     5041 tatagagtct ataggcccac ccccttggct tcgttagaac gcggctacaa ttaatacata
     5101 accttatgta tcatacacat acgatttagg tgacactata gaataacatc cactttgcct
     5161 ttctctccac aggtgtccac tcccaggtcc aactgcacct cggttctatc gaaaacgcgc
     5221 ctgctagcgc caccatgaag accatcattg ccctgagcta catcttctgc ctggtgttcg
     5281 ctgcggccgc caaagctatg catgttgctc agcctgctgt ggtgctggcc tctagcagag
     5341 gaatcgccag ctttgtgtgc gagtacgcct ctcctggaaa ggccactgaa gtgcgcgtga
     5401 cagtgctgag acaggccgat agccaagtga cagaagtgtg cgccgccacc tacatgatgg
     5461 gcaatgagct gaccttcctg gacgacagca tctgtacagg caccagcagc ggcaatcaag
     5521 tgaacctgac aatccagggc ctgagagcca tggacaccgg cctgtatatc tgcaaggtgg
     5581 aactgatgta ccctcctcct tactacctcg gcatcggcaa cggcacccag atctacgtga
     5641 tcgaccctga gccttgtcct gactccgatg gcgcgccgga agatcaagtt gatccaagat
     5701 tgattgatgg caagggcgga tctctggagg tgctgttcca gggaccaggc ggaagcggca
     5761 atcccggcgg cgatctggcc ccccagatgc tgcgcgagct gcaggagacc aatgccgccc
     5821 tgcaggacgt gcgcgagctg ctgcgccacc gcgtgaagga gatcaccttc ctgaagaaca
     5881 ccgtgatgga gtgcgacgcc tgcggtatgc agccctctag aggctccggc tccatcatcc
     5941 cagttgagga ggagaacccg gacttctgga accgcgaggc agccgaggcc ctgggtgccg
     6001 ccaagaagct gcagcctgca cagacagccg ccaagaacct catcatcttc ctgggcgatg
     6061 ggatgggggt gtctacggtg acagctgcca ggatcctaaa agggcagaag aaggacaaac
     6121 tggggcctga gatacccctg gccatggacc gcttcccata tgtggctctg tccaagacat
     6181 acaatgtaga caaacatgtg ccagacagtg gagccacagc cacggcctac ctgtgcgggg
     6241 tcaagggcaa cttccagacc attggcttga gtgcagccgc ccgctttaac cagtgcaaca
     6301 cgacacgcgg caacgaggtc atctccgtga tgaatcgggc caagaaagca gggaagtcag
     6361 tgggagtggt aaccaccaca cgagtgcagc acgcctcgcc agccggcacc tacgcccaca
     6421 cggtgaaccg caactggtac tcggacgccg acgtgcctgc ctcggcccgc caggaggggt
     6481 gccaggacat cgctacgcag ctcatctcca acatggacat tgacgtgatc ctaggtggag
     6541 gccgaaagta catgtttcgc atgggaaccc cagaccctga gtacccagat gactacagcc
     6601 aaggtgggac caggctggac gggaagaatc tggtgcagga atggctggcg aagcgccagg
     6661 gtgcccggta tgtgtggaac cgcactgagc tcatgcaggc ttccctggac ccgtctgtga
     6721 cccatctcat gggtctcttt gagcctggag acatgaaata cgagatccac cgagactcca
     6781 cactggaccc ctccctgatg gagatgacag aggctgccct gcgcctgctg agcaggaacc
     6841 cccgcggctt cttcctcttc gtggagggtg gtcgcatcga ccatggtcat catgaaagca
     6901 gggcttaccg ggcactgact gagacgatca tgttcgacga cgccattgag agggcgggcc
     6961 agctcaccag cgaggaggac acgctgagcc tcgtcactgc cgaccactcc cacgtcttct
     7021 ccttcggagg ctaccccctg cgagggagct ccatcttcgg gctggcccct ggcaaggccc
     7081 gggacaggaa ggcctacacg gtcctcctat acggaaacgg tccaggctat gtgctcaagg
     7141 acggcgcccg gccggatgtt accgagagcg agagcgggag ccccgagtat cggcagcagt
     7201 cagcagtgcc cctggacgaa gagacccacg caggcgagga cgtggcggtg ttcgcgcgcg
     7261 gcccgcaggc gcacctggtt cacggcgtgc aggagcagac cttcatagcg cacgtcatgg
     7321 ccttcgccgc ctgcctggag ccctacaccg cctgcgacct ggcgcccccc gccggcacca
     7381 ccgacgccgc gcacccgggt ggcggaggac tgaacgacat cttcgaggct cagaaaatcg
     7441 aatggcacga acatcaccat caccatcacc atcaccatta aggccggcct gtagtcactg
     7501 gcacaacgcg ttacccggta agccaatcgg g
//
