LOCUS       sequence-417402-        8551 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 8551)
  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 8551)
  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-DLL3-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8551
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        77..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        815..833
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     815..832
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     846..865
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2390..2425
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2435..2458
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             4118..4162
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4163..4189
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(4329..4346)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4335..4446
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4564..4584)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4569..4898
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4749..4884
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4811..4830
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4946..4993
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5012..5611
                     /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(5012..5031)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5508..5528
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5741..5862
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5778..5797)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5832..5851
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5915..6706,6707..6775))
                     /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     6538..6557
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6776..6867)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6909..6928
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6934..6953
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      7039..7627
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7528..7547
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7781..7798
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7896..7913
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7897..7913
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8369..8385
                     /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)."
ORIGIN
        1 ggcattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat
      661 ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc aagagtgacg
      721 taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa cgcggctaca
      781 attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat agaataacat
      841 ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc tcggttctat
      901 cgaaaacgcg cctgctagcg ccaccatgaa gaccatcatt gccctgagct acatcttctg
      961 cctggtgttc gctgcggccg ccgctggtgt ttttgagctg cagatccaca gcttcggccc
     1021 tggacctgga ccaggtgctc ctagatctcc ttgtagcgcc agactgccct gccggctgtt
     1081 cttcagagtg tgtctgaagc ccggcctgag cgaggaagct gctgagtctc cttgtgctct
     1141 gggagccgct ctgtctgcta gaggccctgt gtatacagag cagcctggtg ctcctgctcc
     1201 tgacctgcct cttcctgatg gactgctgca ggtccccttc agagatgcct ggcctggcac
     1261 cttcagcttc atcatcgaga cttggagaga ggaactgggc gaccagatcg gaggacctgc
     1321 ttggagtctg ctggctagag tggccggaag aagaaggctt gctgctggcg gaccttgggc
     1381 cagagatatt caaagagccg gcgcttggga gctgcggttc agctatagag ccagatgcga
     1441 acctcctgcc gtgggcacag catgcaccag actgtgtaga cctagaagcg cccctagcag
     1501 atgtggccca ggactcagac catgtgcacc cctggaagat gagtgcgaag cccctcttgt
     1561 gtgcagagcc ggatgttctc ctgagcacgg cttttgcgaa cagcccggcg agtgcagatg
     1621 tctggaagga tggacaggcc ctctgtgcac cgtgcctgtg tctaccagca gctgtctgag
     1681 ccctagagga ccaagctctg ccacaacagg ctgtctggtg cccggaccag gaccttgtga
     1741 tggcaatcct tgtgccaatg gcggcagctg cagcgagaca cctagatcct tcgagtgcac
     1801 atgccccaga ggcttctacg gcctgagatg tgaagtgtcc ggcgtgacct gtgccgacgg
     1861 accttgtttt aatggcggcc tgtgtgttgg cggcgctgat cctgatagcg cctacatctg
     1921 tcactgccct ccaggcttcc agggcagcaa ctgcgagaag cgagtggaca gatgttccct
     1981 gcagccttgc agaaacggcg gactgtgtct ggatctggga cacgccctga gatgcagatg
     2041 tagagccggc tttgccggac ctagatgcga gcacgatctg gacgattgtg ccggcagagc
     2101 ctgcgctaat ggcggaacat gtgttgaagg cggcggagcc cacagatgct cttgtgccct
     2161 tggctttggc ggcagagact gcagagaaag agccgatcct tgcgccgcca gaccttgcgc
     2221 tcacggtgga agatgttacg cccactttag cggcctcgtg tgcgcttgtg ccccaggata
     2281 tatgggcgcc agatgtgaat tcccagtgca ccctgatggc gcctctgcac ttcctgctgc
     2341 tccacctgga cttagacctg gcgaccctca gagatacctt ggcgcgccgg aagatcaagt
     2401 tgatccaaga ttgattgatg gcaagggcgg atctctggag gtgctgttcc agggaccagg
     2461 cggaagcggc aatcccggcg gcgatctggc cccccagatg ctgcgcgagc tgcaggagac
     2521 caatgccgcc ctgcaggacg tgcgcgagct gctgcgccac cgcgtgaagg agatcacctt
     2581 cctgaagaac accgtgatgg agtgcgacgc ctgcggtatg cagccctcta gaggctccgg
     2641 ctccatcatc ccagttgagg aggagaaccc ggacttctgg aaccgcgagg cagccgaggc
     2701 cctgggtgcc gccaagaagc tgcagcctgc acagacagcc gccaagaacc tcatcatctt
     2761 cctgggcgat gggatggggg tgtctacggt gacagctgcc aggatcctaa aagggcagaa
     2821 gaaggacaaa ctggggcctg agatacccct ggccatggac cgcttcccat atgtggctct
     2881 gtccaagaca tacaatgtag acaaacatgt gccagacagt ggagccacag ccacggccta
     2941 cctgtgcggg gtcaagggca acttccagac cattggcttg agtgcagccg cccgctttaa
     3001 ccagtgcaac acgacacgcg gcaacgaggt catctccgtg atgaatcggg ccaagaaagc
     3061 agggaagtca gtgggagtgg taaccaccac acgagtgcag cacgcctcgc cagccggcac
     3121 ctacgcccac acggtgaacc gcaactggta ctcggacgcc gacgtgcctg cctcggcccg
     3181 ccaggagggg tgccaggaca tcgctacgca gctcatctcc aacatggaca ttgacgtgat
     3241 cctaggtgga ggccgaaagt acatgtttcg catgggaacc ccagaccctg agtacccaga
     3301 tgactacagc caaggtggga ccaggctgga cgggaagaat ctggtgcagg aatggctggc
     3361 gaagcgccag ggtgcccggt atgtgtggaa ccgcactgag ctcatgcagg cttccctgga
     3421 cccgtctgtg acccatctca tgggtctctt tgagcctgga gacatgaaat acgagatcca
     3481 ccgagactcc acactggacc cctccctgat ggagatgaca gaggctgccc tgcgcctgct
     3541 gagcaggaac ccccgcggct tcttcctctt cgtggagggt ggtcgcatcg accatggtca
     3601 tcatgaaagc agggcttacc gggcactgac tgagacgatc atgttcgacg acgccattga
     3661 gagggcgggc cagctcacca gcgaggagga cacgctgagc ctcgtcactg ccgaccactc
     3721 ccacgtcttc tccttcggag gctaccccct gcgagggagc tccatcttcg ggctggcccc
     3781 tggcaaggcc cgggacagga aggcctacac ggtcctccta tacggaaacg gtccaggcta
     3841 tgtgctcaag gacggcgccc ggccggatgt taccgagagc gagagcggga gccccgagta
     3901 tcggcagcag tcagcagtgc ccctggacga agagacccac gcaggcgagg acgtggcggt
     3961 gttcgcgcgc ggcccgcagg cgcacctggt tcacggcgtg caggagcaga ccttcatagc
     4021 gcacgtcatg gccttcgccg cctgcctgga gccctacacc gcctgcgacc tggcgccccc
     4081 cgccggcacc accgacgccg cgcacccggg tggcggagga ctgaacgaca tcttcgaggc
     4141 tcagaaaatc gaatggcacg aacatcacca tcaccatcac catcaccatt aaggccggcc
     4201 tgtagtcact ggcacaacgc gttacccggt aagccaatcg ggtatacacg gtcgtcatac
     4261 tgcagacagg gttcttctac tttgcaagat agtcttgagt agtaaaataa atagatagag
     4321 aaaatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     4381 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     4441 tgcatcacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     4501 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     4561 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     4621 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     4681 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     4741 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     4801 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     4861 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     4921 ccattttcgg atctgatcag cacgtgttga caattaatca tcggcatagt atatcggcat
     4981 agtataatac gacaaggtga ggaactaaac catgaccgag tacaagccca cggtgcgcct
     5041 cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta
     5101 ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca
     5161 agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg
     5221 cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga
     5281 gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga
     5341 aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt
     5401 ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc
     5461 ggccgagcgc gccggggtgc ccgccttcct ggagacatcc gcgccccgca acctcccctt
     5521 ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac
     5581 ctggtgcatg acccgcaagc ccggtgcctg acacgtgcta cgagatttcg attccaccgc
     5641 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     5701 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     5761 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     5821 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     5881 gacctctagc tagagcttgg cgtaatcatg gtcattacca atgcttaatc agtgaggcac
     5941 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     6001 taactacgat acgggagggc ttaccatctg gccccagcgc tgcgatgata ccgcgagaac
     6061 cacgctcacc ggctccggat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     6121 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6181 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccatcgct acaggcatcg
     6241 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6301 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6361 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6421 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6481 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6541 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6601 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6661 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6721 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatattct
     6781 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6841 ttgaatgtat ttagaaaaat aaacaaatag gggtcagtgt tacaaccaat taaccaattc
     6901 tgaacattat cgcgagccca tttatacctg aatatggctc ataacacccc ttgctcatga
     6961 ccaaaatccc ttaacgtgag ttacgcgcgc gtcgttccac tgagcgtcag accccgtaga
     7021 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     7081 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     7141 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     7201 gtagttagcc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     7261 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     7321 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     7381 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     7441 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     7501 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     7561 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     7621 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     7681 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     7741 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     7801 agcggaaggc gagagtaggg aactgccagg catcaaacta agcagaaggc ccctgacgga
     7861 tggccttttt gcgtttctac aaactctttc tgtgttgtaa aacgacggcc agtcttaagc
     7921 tcgggccccc tgggcggttc tgataacgag taatcgttaa tccgcaaata acgtaaaaac
     7981 ccgcttcggc gggttttttt atggggggag tttagggaaa gagcatttgt cagaatattt
     8041 aagggcgcct gtcactttgc ttgatatatg agaattattt aaccttataa atgagaaaaa
     8101 agcaacgcac tttaaataag atacgttgct ttttcgattg atgaacacct ataattaaac
     8161 tattcatcta ttatttatga ttttttgtat atacaatatt tctagtttgt taaagagaat
     8221 taagaaaata aatctcgaaa ataataaagg gaaaatcagt ttttgatatc aaaattatac
     8281 atgtcaacga taatacaaaa tataatacaa actataagat gttatcagta tttattatca
     8341 tttagaataa attttgtgtc gcccttaatt gtgagcggat aacaattacg agcttcatgc
     8401 acagtggcgt tgactcaata ttggccatta gccatattat tcattggtta tatagcataa
     8461 atcaatattg gctattggcc attgcatacg ttgtatctat atcataatat gtacatttat
     8521 attggctcat gtccaatatg accgccatgt t
//
