LOCUS       sequence-303281-        7549 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 7549)
  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 7549)
  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-SCN4B-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7549
                     /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             3154..3189
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             3199..3222
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             4882..4926
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4927..4953
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(5093..5110)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5099..5210
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5328..5348)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5333..5662
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5513..5648
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5575..5594
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5710..5757
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5776..6375
                     /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(5776..5795)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6272..6292
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6505..6626
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6542..6561)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6596..6615
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6679..7470,7471..7539))
                     /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     7302..7321
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7540..7549,1..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 ggaagtgtct
     2761 gtgggcaaag ccacagacat ctacgccgtg aacggcaccg agatcctgct gccttgtacc
     2821 ttctccagct gcttcggctt cgaggacctg cacttcagat ggacctacaa cagcagcgac
     2881 gccttcaaga tcctgatcga gggcaccgtg aagaacgaga agtctgaccc caaagtgacc
     2941 ctgaaggacg acgacagaat caccctcgtg ggcagcacca aagaaaagat gaacaacatc
     3001 agcatcgtgc tgcgcgacct ggaattcagc gacaccggca agtacacctg tcacgtgaag
     3061 aatcccaaag agaacaacct gcagcaccac gccaccatct ttctgcaagt ggtggacaga
     3121 ctggaagagg tggacaacac cgttggcgcg ccggaagatc aagttgatcc aagattgatt
     3181 gatggcaagg gcggatctct ggaggtgctg ttccagggac caggcggaag cggcaatccc
     3241 ggcggcgatc tggcccccca gatgctgcgc gagctgcagg agaccaatgc cgccctgcag
     3301 gacgtgcgcg agctgctgcg ccaccgcgtg aaggagatca ccttcctgaa gaacaccgtg
     3361 atggagtgcg acgcctgcgg tatgcagccc tctagaggct ccggctccat catcccagtt
     3421 gaggaggaga acccggactt ctggaaccgc gaggcagccg aggccctggg tgccgccaag
     3481 aagctgcagc ctgcacagac agccgccaag aacctcatca tcttcctggg cgatgggatg
     3541 ggggtgtcta cggtgacagc tgccaggatc ctaaaagggc agaagaagga caaactgggg
     3601 cctgagatac ccctggccat ggaccgcttc ccatatgtgg ctctgtccaa gacatacaat
     3661 gtagacaaac atgtgccaga cagtggagcc acagccacgg cctacctgtg cggggtcaag
     3721 ggcaacttcc agaccattgg cttgagtgca gccgcccgct ttaaccagtg caacacgaca
     3781 cgcggcaacg aggtcatctc cgtgatgaat cgggccaaga aagcagggaa gtcagtggga
     3841 gtggtaacca ccacacgagt gcagcacgcc tcgccagccg gcacctacgc ccacacggtg
     3901 aaccgcaact ggtactcgga cgccgacgtg cctgcctcgg cccgccagga ggggtgccag
     3961 gacatcgcta cgcagctcat ctccaacatg gacattgacg tgatcctagg tggaggccga
     4021 aagtacatgt ttcgcatggg aaccccagac cctgagtacc cagatgacta cagccaaggt
     4081 gggaccaggc tggacgggaa gaatctggtg caggaatggc tggcgaagcg ccagggtgcc
     4141 cggtatgtgt ggaaccgcac tgagctcatg caggcttccc tggacccgtc tgtgacccat
     4201 ctcatgggtc tctttgagcc tggagacatg aaatacgaga tccaccgaga ctccacactg
     4261 gacccctccc tgatggagat gacagaggct gccctgcgcc tgctgagcag gaacccccgc
     4321 ggcttcttcc tcttcgtgga gggtggtcgc atcgaccatg gtcatcatga aagcagggct
     4381 taccgggcac tgactgagac gatcatgttc gacgacgcca ttgagagggc gggccagctc
     4441 accagcgagg aggacacgct gagcctcgtc actgccgacc actcccacgt cttctccttc
     4501 ggaggctacc ccctgcgagg gagctccatc ttcgggctgg cccctggcaa ggcccgggac
     4561 aggaaggcct acacggtcct cctatacgga aacggtccag gctatgtgct caaggacggc
     4621 gcccggccgg atgttaccga gagcgagagc gggagccccg agtatcggca gcagtcagca
     4681 gtgcccctgg acgaagagac ccacgcaggc gaggacgtgg cggtgttcgc gcgcggcccg
     4741 caggcgcacc tggttcacgg cgtgcaggag cagaccttca tagcgcacgt catggccttc
     4801 gccgcctgcc tggagcccta caccgcctgc gacctggcgc cccccgccgg caccaccgac
     4861 gccgcgcacc cgggtggcgg aggactgaac gacatcttcg aggctcagaa aatcgaatgg
     4921 cacgaacatc accatcacca tcaccatcac cattaaggcc ggcctgtagt cactggcaca
     4981 acgcgttacc cggtaagcca atcgggtata cacggtcgtc atactgcaga cagggttctt
     5041 ctactttgca agatagtctt gagtagtaaa ataaatagat agagaaaatc agcctcgact
     5101 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     5161 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc acaacactca
     5221 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     5281 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     5341 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     5401 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     5461 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     5521 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     5581 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     5641 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     5701 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     5761 gtgaggaact aaaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     5821 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     5881 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     5941 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     6001 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     6061 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     6121 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     6181 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     6241 gtgcccgcct tcctggagac atccgcgccc cgcaacctcc ccttctacga gcggctcggc
     6301 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     6361 aagcccggtg cctgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg
     6421 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     6481 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     6541 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     6601 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     6661 ttggcgtaat catggtcatt accaatgctt aatcagtgag gcacctatct cagcgatctg
     6721 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     6781 gggcttacca tctggcccca gcgctgcgat gataccgcga gaaccacgct caccggctcc
     6841 ggatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     6901 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     6961 agttaatagt ttgcgcaacg ttgttgccat cgctacaggc atcgtggtgt cacgctcgtc
     7021 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     7081 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     7141 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     7201 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     7261 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     7321 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     7381 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     7441 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     7501 aaagggaata agggcgacac ggaaatgttg aatactcata ttcttcctt
//
