LOCUS       sequence-491726-        7966 bp    DNA     circular SYN 04-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 7966)
  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 7966)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 4, 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-IZUMO1-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7966
                     /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             3616..3639
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5299..5343
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5344..5370
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(5510..5527)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5516..5627
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5745..5765)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5750..6079
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5930..6065
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5992..6011
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6127..6174
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6193..6792
                     /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(6193..6212)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6689..6709
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6922..7043
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6959..6978)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7013..7032
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7096..7887,7888..7956))
                     /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     7719..7738
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7957..7966,1..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 tgtgatctgt
     2761 gatccttctg tggtgctggc cctgaagtcc ctggaaaagg attatctgcc tggccacctg
     2821 gacgccaagc accacaaagc catgatggaa cgcgtggaaa acgccgtgaa ggacttccaa
     2881 gagctgagcc tgaacgagga cgcctacatg ggagttgtgg atgaggccac actgcagaaa
     2941 ggcagctggt ccctgctgaa ggacctgaag agaatcaccg acagcgacgt gaagggcgac
     3001 ctgttcgtga aagaactgtt ctggatgctg cacctccaga aagagacatt cgccacctac
     3061 gtggcccggt ttcagaaaga ggcctactgt cctaacaagt gcggcgtgat gctgcagacc
     3121 ctgatctggt gcaagaactg caagaaagag gtgcacgcct gccggaagtc ctacgactgt
     3181 ggcgagagaa acgtggaagt gccccagatg gaagatatga tcctggactg cgagctgaac
     3241 tggcaccagg cctctgaggg cctgaccgac tacagctttt acagagtgtg gggcaacaac
     3301 accgagacac tggtgtccaa gggcaaagag gccactctga ccaagcctat ggttggacct
     3361 gaggacgccg gcagctatag atgtgaactg ggctccgtga atagcagccc cgccaccatc
     3421 atcaatttcc acgtgaccgt gctgcccaag atgatcaaag aggaaaagcc ctctcctaac
     3481 atcgtgaccc ctggcgaggc cacaacagag agcagtattt ctctgcagcc cctgcagcct
     3541 gagaagatgc tggctagtag aggcgcgccg gaagatcaag ttgatccaag attgattgat
     3601 ggcaagggcg gatctctgga ggtgctgttc cagggaccag gcggaagcgg caatcccggc
     3661 ggcgatctgg ccccccagat gctgcgcgag ctgcaggaga ccaatgccgc cctgcaggac
     3721 gtgcgcgagc tgctgcgcca ccgcgtgaag gagatcacct tcctgaagaa caccgtgatg
     3781 gagtgcgacg cctgcggtat gcagccctct agaggctccg gctccatcat cccagttgag
     3841 gaggagaacc cggacttctg gaaccgcgag gcagccgagg ccctgggtgc cgccaagaag
     3901 ctgcagcctg cacagacagc cgccaagaac ctcatcatct tcctgggcga tgggatgggg
     3961 gtgtctacgg tgacagctgc caggatccta aaagggcaga agaaggacaa actggggcct
     4021 gagatacccc tggccatgga ccgcttccca tatgtggctc tgtccaagac atacaatgta
     4081 gacaaacatg tgccagacag tggagccaca gccacggcct acctgtgcgg ggtcaagggc
     4141 aacttccaga ccattggctt gagtgcagcc gcccgcttta accagtgcaa cacgacacgc
     4201 ggcaacgagg tcatctccgt gatgaatcgg gccaagaaag cagggaagtc agtgggagtg
     4261 gtaaccacca cacgagtgca gcacgcctcg ccagccggca cctacgccca cacggtgaac
     4321 cgcaactggt actcggacgc cgacgtgcct gcctcggccc gccaggaggg gtgccaggac
     4381 atcgctacgc agctcatctc caacatggac attgacgtga tcctaggtgg aggccgaaag
     4441 tacatgtttc gcatgggaac cccagaccct gagtacccag atgactacag ccaaggtggg
     4501 accaggctgg acgggaagaa tctggtgcag gaatggctgg cgaagcgcca gggtgcccgg
     4561 tatgtgtgga accgcactga gctcatgcag gcttccctgg acccgtctgt gacccatctc
     4621 atgggtctct ttgagcctgg agacatgaaa tacgagatcc accgagactc cacactggac
     4681 ccctccctga tggagatgac agaggctgcc ctgcgcctgc tgagcaggaa cccccgcggc
     4741 ttcttcctct tcgtggaggg tggtcgcatc gaccatggtc atcatgaaag cagggcttac
     4801 cgggcactga ctgagacgat catgttcgac gacgccattg agagggcggg ccagctcacc
     4861 agcgaggagg acacgctgag cctcgtcact gccgaccact cccacgtctt ctccttcgga
     4921 ggctaccccc tgcgagggag ctccatcttc gggctggccc ctggcaaggc ccgggacagg
     4981 aaggcctaca cggtcctcct atacggaaac ggtccaggct atgtgctcaa ggacggcgcc
     5041 cggccggatg ttaccgagag cgagagcggg agccccgagt atcggcagca gtcagcagtg
     5101 cccctggacg aagagaccca cgcaggcgag gacgtggcgg tgttcgcgcg cggcccgcag
     5161 gcgcacctgg ttcacggcgt gcaggagcag accttcatag cgcacgtcat ggccttcgcc
     5221 gcctgcctgg agccctacac cgcctgcgac ctggcgcccc ccgccggcac caccgacgcc
     5281 gcgcacccgg gtggcggagg actgaacgac atcttcgagg ctcagaaaat cgaatggcac
     5341 gaacatcacc atcaccatca ccatcaccat taaggccggc ctgtagtcac tggcacaacg
     5401 cgttacccgg taagccaatc gggtatacac ggtcgtcata ctgcagacag ggttcttcta
     5461 ctttgcaaga tagtcttgag tagtaaaata aatagataga gaaaatcagc ctcgactgtg
     5521 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     5581 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcaca acactcaacc
     5641 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     5701 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt
     5761 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     5821 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     5881 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     5941 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     6001 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg
     6061 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca
     6121 gcacgtgttg acaattaatc atcggcatag tatatcggca tagtataata cgacaaggtg
     6181 aggaactaaa ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     6241 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     6301 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     6361 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     6421 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     6481 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac
     6541 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc
     6601 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg
     6661 cccgccttcc tggagacatc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     6721 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     6781 cccggtgcct gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
     6841 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     6901 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     6961 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     7021 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     7081 gcgtaatcat ggtcattacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     7141 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     7201 cttaccatct ggccccagcg ctgcgatgat accgcgagaa ccacgctcac cggctccgga
     7261 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     7321 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     7381 taatagtttg cgcaacgttg ttgccatcgc tacaggcatc gtggtgtcac gctcgtcgtt
     7441 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     7501 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     7561 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     7621 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     7681 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     7741 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     7801 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7861 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7921 gggaataagg gcgacacgga aatgttgaat actcatattc ttcctt
//
