LOCUS       sequence-466830-        9046 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 9046)
  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 9046)
  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-SEMA6A-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..9046
                     /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             4651..4686
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4696..4719
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             6379..6423
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6424..6450
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6590..6607)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6596..6707
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6825..6845)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6830..7159
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7010..7145
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7072..7091
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7207..7254
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7273..7872
                     /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(7273..7292)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7769..7789
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    8002..8123
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8039..8058)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8093..8112
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8176..8967,8968..9036))
                     /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     8799..8818
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(9037..9046,1..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 cggccgccgg atttcctgag
     2761 gatagcgagc ctatcagcat cagccacggc aactacacca agcagtaccc cgtgttcgtg
     2821 ggccacaagc ctggcagaaa caccacacag agacaccggc tggacatcca gatgatcatg
     2881 attatgaacg gcaccctgta tatcgccgcc agggaccaca tctacaccgt ggacatcgat
     2941 accagccaca ccgaggaaat ctactgcagc aagaagctga cctggaagtc cagacaggcc
     3001 gacgtggaca cctgtcggat gaagggcaag cacaaggacg agtgccacaa cttcatcaag
     3061 gtgctgctga agaagaacga cgacgccctg ttcgtgtgcg gcaccaacgc cttcaatccc
     3121 agctgccgga actacaagat ggacaccctg gaacctttcg gcgacgagtt cagcggcatg
     3181 gccagatgtc cttacgatgc caagcacgcc aacgtggccc tgtttgccga tggcaagctg
     3241 tactctgcta ccgtgaccga cttcctggcc atcgacgccg tgatctacag aagcctgggc
     3301 gagagcccta cactgagaac cgtgaagcac gacagcaagt ggctgaaaga accctacttt
     3361 gtgcaggccg tggactacgg cgactacatc tactttttct tccgggaaat cgccgtcgag
     3421 tacaacacca tgggcaaagt ggtgttcccc agagtggccc aagtgtgcaa gaacgatatg
     3481 ggcggctctc agcgcgtgct ggaaaagcag tggaccagct tcctgaaggc ccggctgaat
     3541 tgctctgtgc ctggcgacag ccacttctac ttcaacatcc tgcaggctgt gaccgacgtg
     3601 atcagaatca acggcagaga tgtggtgctg gccaccttca gcacccctta caatagcatc
     3661 cctggcagcg ccgtgtgcgc ctacgatatg ctggatatcg ccagcgtgtt caccggcaga
     3721 ttcaaagagc agaagtcccc tgacagcacc tggacaccag tgcctgatga gagagtgccc
     3781 aagcctagac ctggctgttg tgccggatct agcagcctgg aaagatacgc caccagcaac
     3841 gagttccccg acgacaccct gaatttcatc aagacacacc ctctgatgga cgaggccgtg
     3901 cctagcatct tcaacagacc ctggttcctg agaaccatgg tccgataccg gctgaccaag
     3961 atcgccgtgg atacagccgc tggaccctac cagaatcaca ccgtggtctt tctgggcagc
     4021 gagaagggaa tcatcctgaa gtttctggcc cggatcggca acagcggctt tctgaacgat
     4081 agcctgttcc tggaagagat gagcgtgtac aactccgaga agtgcagcta cgacggcgtg
     4141 gaagataagc ggatcatggg catgcagctg gacagagcca gcagctctct gtacgtggcc
     4201 ttctccacct gtgtgattaa ggtgcccctg ggcagatgcg agagatacgg caagtgcaag
     4261 aaaacctgca ttgccagcag ggacccctac tgcggctgga tcaaagaagg cggagcctgc
     4321 tctcatctga gccccaatag cagactgacc ttcgagcagg acatcgagag aggcaatacc
     4381 gacggcctgg gagactgcca caattccttt gtggccctga acggccactc aagcagcctg
     4441 ctgcctagca ccacaaccag cgatagcaca gcccaagagg gctacgaaag cagaggcggc
     4501 atgctggact ggaagcacct cctggatagc cccgatagca ccgatcctct gggagccgtg
     4561 tctagccaca accaccagga caagaagggc gtcatcagag agagctacct gaagggccac
     4621 gatcagctgg tgcctgtgac aggcgcgccg gaagatcaag ttgatccaag attgattgat
     4681 ggcaagggcg gatctctgga ggtgctgttc cagggaccag gcggaagcgg caatcccggc
     4741 ggcgatctgg ccccccagat gctgcgcgag ctgcaggaga ccaatgccgc cctgcaggac
     4801 gtgcgcgagc tgctgcgcca ccgcgtgaag gagatcacct tcctgaagaa caccgtgatg
     4861 gagtgcgacg cctgcggtat gcagccctct agaggctccg gctccatcat cccagttgag
     4921 gaggagaacc cggacttctg gaaccgcgag gcagccgagg ccctgggtgc cgccaagaag
     4981 ctgcagcctg cacagacagc cgccaagaac ctcatcatct tcctgggcga tgggatgggg
     5041 gtgtctacgg tgacagctgc caggatccta aaagggcaga agaaggacaa actggggcct
     5101 gagatacccc tggccatgga ccgcttccca tatgtggctc tgtccaagac atacaatgta
     5161 gacaaacatg tgccagacag tggagccaca gccacggcct acctgtgcgg ggtcaagggc
     5221 aacttccaga ccattggctt gagtgcagcc gcccgcttta accagtgcaa cacgacacgc
     5281 ggcaacgagg tcatctccgt gatgaatcgg gccaagaaag cagggaagtc agtgggagtg
     5341 gtaaccacca cacgagtgca gcacgcctcg ccagccggca cctacgccca cacggtgaac
     5401 cgcaactggt actcggacgc cgacgtgcct gcctcggccc gccaggaggg gtgccaggac
     5461 atcgctacgc agctcatctc caacatggac attgacgtga tcctaggtgg aggccgaaag
     5521 tacatgtttc gcatgggaac cccagaccct gagtacccag atgactacag ccaaggtggg
     5581 accaggctgg acgggaagaa tctggtgcag gaatggctgg cgaagcgcca gggtgcccgg
     5641 tatgtgtgga accgcactga gctcatgcag gcttccctgg acccgtctgt gacccatctc
     5701 atgggtctct ttgagcctgg agacatgaaa tacgagatcc accgagactc cacactggac
     5761 ccctccctga tggagatgac agaggctgcc ctgcgcctgc tgagcaggaa cccccgcggc
     5821 ttcttcctct tcgtggaggg tggtcgcatc gaccatggtc atcatgaaag cagggcttac
     5881 cgggcactga ctgagacgat catgttcgac gacgccattg agagggcggg ccagctcacc
     5941 agcgaggagg acacgctgag cctcgtcact gccgaccact cccacgtctt ctccttcgga
     6001 ggctaccccc tgcgagggag ctccatcttc gggctggccc ctggcaaggc ccgggacagg
     6061 aaggcctaca cggtcctcct atacggaaac ggtccaggct atgtgctcaa ggacggcgcc
     6121 cggccggatg ttaccgagag cgagagcggg agccccgagt atcggcagca gtcagcagtg
     6181 cccctggacg aagagaccca cgcaggcgag gacgtggcgg tgttcgcgcg cggcccgcag
     6241 gcgcacctgg ttcacggcgt gcaggagcag accttcatag cgcacgtcat ggccttcgcc
     6301 gcctgcctgg agccctacac cgcctgcgac ctggcgcccc ccgccggcac caccgacgcc
     6361 gcgcacccgg gtggcggagg actgaacgac atcttcgagg ctcagaaaat cgaatggcac
     6421 gaacatcacc atcaccatca ccatcaccat taaggccggc ctgtagtcac tggcacaacg
     6481 cgttacccgg taagccaatc gggtatacac ggtcgtcata ctgcagacag ggttcttcta
     6541 ctttgcaaga tagtcttgag tagtaaaata aatagataga gaaaatcagc ctcgactgtg
     6601 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     6661 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcaca acactcaacc
     6721 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     6781 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt
     6841 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     6901 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     6961 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     7021 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     7081 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg
     7141 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca
     7201 gcacgtgttg acaattaatc atcggcatag tatatcggca tagtataata cgacaaggtg
     7261 aggaactaaa ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     7321 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     7381 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     7441 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     7501 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     7561 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac
     7621 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc
     7681 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg
     7741 cccgccttcc tggagacatc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     7801 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     7861 cccggtgcct gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
     7921 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     7981 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     8041 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     8101 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     8161 gcgtaatcat ggtcattacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     8221 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     8281 cttaccatct ggccccagcg ctgcgatgat accgcgagaa ccacgctcac cggctccgga
     8341 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     8401 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     8461 taatagtttg cgcaacgttg ttgccatcgc tacaggcatc gtggtgtcac gctcgtcgtt
     8521 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     8581 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     8641 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     8701 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     8761 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     8821 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     8881 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     8941 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     9001 gggaataagg gcgacacgga aatgttgaat actcatattc ttcctt
//
