LOCUS       sequence-303999-        8623 bp    DNA     circular SYN 15-DEC-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 8623)
  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 8623)
  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-IGSF1-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8623
                     /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             4228..4263
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4273..4296
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5956..6000
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6001..6027
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6167..6184)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6173..6284
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6402..6422)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6407..6736
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6587..6722
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6649..6668
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6784..6831
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6850..7449
                     /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(6850..6869)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7346..7366
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7579..7700
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7616..7635)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7670..7689
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7753..8544,8545..8613))
                     /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     8376..8395
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8614..8623,1..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 cggccgccat gacctctatt
     2761 gtgatggacc ctcagcctga gctgtggatc gagagcaatt accctcaggc tccctgggag
     2821 aacatcaccc tgtggtgtag aagccccagc cggatcagca gcaagttcct gctgctgaag
     2881 gacaagaccc agatgacctg gatcagaccc agccacaaga ccttccaggt gtcctttctg
     2941 atcggcgccc tgacagagag caatgccggc ctgtacagat gctgctactg gaaagagaca
     3001 ggctggtcca agcctagcaa ggtgctggaa ctggaagctc ctggacagct gcccaagcct
     3061 atcttctgga ttcaagccga gacacccgct ctgcccggct gcaatgtgaa tatcctgtgt
     3121 cacggctggc tgcaggacct ggtgttcatg ctgttcaaag agggctacgc cgagcctgtg
     3181 gattaccagg tgccaacagg caccatggcc atcttcagca tcgacaatct gacccctgag
     3241 gacgagggcg tgtacatctg tagaacccac atccagatgc tccccacact ttggagcgag
     3301 cccagcaatc ctctgaaact ggtggtggcc ggactgtacc ccaagccaac acttacagcc
     3361 catcctggac ctatcatggc ccctggcgag agcctgaatc tgagatgcca gggacctatc
     3421 tacggcatga cattcgccct gatgagagtc gaggacctgg aaaagagctt ctaccacaag
     3481 aaaaccatca agaacgaggc caacttcttc tttcaatccc tgaaaatcca ggacaccggc
     3541 cactacctgt gcttctacta cgatgccagc tacagaggca gcctgctgag cgacgtgctg
     3601 aaaatctggg tcaccgacac attccccaag acctggctgc tggctagacc ttctgctgtg
     3661 gtgcagatgg gccagaacgt gtccctgaga tgtagaggcc ctgtggatgg tgtcggactg
     3721 gccctgtata agaaaggcga ggacaagccc ctgcagttcc tggatgccac ctccatcgac
     3781 gacaatacca gcttcttcct gaacaacgtg acctacagcg acaccggcat ctacagctgt
     3841 cactacctgc tgacatggaa aaccagcatc agaatgccta gccacaacac cgtggaactg
     3901 atggtggtgg acaagcctcc taagccttct ctgagcgcct ggcctagcac agtgttcaag
     3961 ctgggcaaag ccatcactct gcagtgtaga gtgtctcacc ccgtgctgga attcagcctg
     4021 gaatgggaag agagggaaac cttccagaaa ttcagcgtga acggcgactt catcatcagc
     4081 aacgtggacg gcaaaggcac cggcacctac tcatgctcct acagagtgga aacacacccc
     4141 aacatctggt cccacagaag cgagcctctg aagctgatgg gacctgccgg ctacctgacc
     4201 tggaactatg tgctgaatgg cgcgccggaa gatcaagttg atccaagatt gattgatggc
     4261 aagggcggat ctctggaggt gctgttccag ggaccaggcg gaagcggcaa tcccggcggc
     4321 gatctggccc cccagatgct gcgcgagctg caggagacca atgccgccct gcaggacgtg
     4381 cgcgagctgc tgcgccaccg cgtgaaggag atcaccttcc tgaagaacac cgtgatggag
     4441 tgcgacgcct gcggtatgca gccctctaga ggctccggct ccatcatccc agttgaggag
     4501 gagaacccgg acttctggaa ccgcgaggca gccgaggccc tgggtgccgc caagaagctg
     4561 cagcctgcac agacagccgc caagaacctc atcatcttcc tgggcgatgg gatgggggtg
     4621 tctacggtga cagctgccag gatcctaaaa gggcagaaga aggacaaact ggggcctgag
     4681 atacccctgg ccatggaccg cttcccatat gtggctctgt ccaagacata caatgtagac
     4741 aaacatgtgc cagacagtgg agccacagcc acggcctacc tgtgcggggt caagggcaac
     4801 ttccagacca ttggcttgag tgcagccgcc cgctttaacc agtgcaacac gacacgcggc
     4861 aacgaggtca tctccgtgat gaatcgggcc aagaaagcag ggaagtcagt gggagtggta
     4921 accaccacac gagtgcagca cgcctcgcca gccggcacct acgcccacac ggtgaaccgc
     4981 aactggtact cggacgccga cgtgcctgcc tcggcccgcc aggaggggtg ccaggacatc
     5041 gctacgcagc tcatctccaa catggacatt gacgtgatcc taggtggagg ccgaaagtac
     5101 atgtttcgca tgggaacccc agaccctgag tacccagatg actacagcca aggtgggacc
     5161 aggctggacg ggaagaatct ggtgcaggaa tggctggcga agcgccaggg tgcccggtat
     5221 gtgtggaacc gcactgagct catgcaggct tccctggacc cgtctgtgac ccatctcatg
     5281 ggtctctttg agcctggaga catgaaatac gagatccacc gagactccac actggacccc
     5341 tccctgatgg agatgacaga ggctgccctg cgcctgctga gcaggaaccc ccgcggcttc
     5401 ttcctcttcg tggagggtgg tcgcatcgac catggtcatc atgaaagcag ggcttaccgg
     5461 gcactgactg agacgatcat gttcgacgac gccattgaga gggcgggcca gctcaccagc
     5521 gaggaggaca cgctgagcct cgtcactgcc gaccactccc acgtcttctc cttcggaggc
     5581 taccccctgc gagggagctc catcttcggg ctggcccctg gcaaggcccg ggacaggaag
     5641 gcctacacgg tcctcctata cggaaacggt ccaggctatg tgctcaagga cggcgcccgg
     5701 ccggatgtta ccgagagcga gagcgggagc cccgagtatc ggcagcagtc agcagtgccc
     5761 ctggacgaag agacccacgc aggcgaggac gtggcggtgt tcgcgcgcgg cccgcaggcg
     5821 cacctggttc acggcgtgca ggagcagacc ttcatagcgc acgtcatggc cttcgccgcc
     5881 tgcctggagc cctacaccgc ctgcgacctg gcgccccccg ccggcaccac cgacgccgcg
     5941 cacccgggtg gcggaggact gaacgacatc ttcgaggctc agaaaatcga atggcacgaa
     6001 catcaccatc accatcacca tcaccattaa ggccggcctg tagtcactgg cacaacgcgt
     6061 tacccggtaa gccaatcggg tatacacggt cgtcatactg cagacagggt tcttctactt
     6121 tgcaagatag tcttgagtag taaaataaat agatagagaa aatcagcctc gactgtgcct
     6181 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     6241 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcacaaca ctcaacccta
     6301 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     6361 atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg
     6421 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     6481 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     6541 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     6601 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     6661 ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     6721 cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcagca
     6781 cgtgttgaca attaatcatc ggcatagtat atcggcatag tataatacga caaggtgagg
     6841 aactaaacca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
     6901 agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
     6961 gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
     7021 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc
     7081 acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag
     7141 ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg
     7201 cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag
     7261 ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc
     7321 gccttcctgg agacatccgc gccccgcaac ctccccttct acgagcggct cggcttcacc
     7381 gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc
     7441 ggtgcctgac acgtgctacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     7501 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     7561 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     7621 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     7681 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     7741 taatcatggt cattaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     7801 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     7861 accatctggc cccagcgctg cgatgatacc gcgagaacca cgctcaccgg ctccggattt
     7921 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     7981 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     8041 tagtttgcgc aacgttgttg ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     8101 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     8161 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     8221 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     8281 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     8341 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     8401 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     8461 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     8521 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     8581 aataagggcg acacggaaat gttgaatact catattcttc ctt
//
