LOCUS       sequence-421006-        7774 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 7774)
  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 7774)
  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-TNFSF12-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7774
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             42..68
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(208..225)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    214..325
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(443..463)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        448..777
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      628..763
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     690..709
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        825..872
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             891..1490
                     /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(891..910)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1387..1407
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1620..1741
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1657..1676)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1711..1730
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(1794..2585,2586..2654))
                     /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     2417..2436
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2655..2746)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2788..2807
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2813..2832
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2918..3506
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3407..3426
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3660..3677
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     3775..3792
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3776..3792
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4248..4264
                     /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        4431..4810
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4811..5014
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4965..4985
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5011..5035
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        5245..5263
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     5245..5262
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     5276..5295
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             6043..6078
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             6088..6111
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(7771..7774,1..41)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
ORIGIN
        1 tgaacgacat cttcgaggct cagaaaatcg aatggcacga acatcaccat caccatcacc
       61 atcaccatta aggccggcct gtagtcactg gcacaacgcg ttacccggta agccaatcgg
      121 gtatacacgg tcgtcatact gcagacaggg ttcttctact ttgcaagata gtcttgagta
      181 gtaaaataaa tagatagaga aaatcagcct cgactgtgcc ttctagttgc cagccatctg
      241 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
      301 cctaataaaa tgaggaaatt gcatcacaac actcaaccct atctcggtct attcttttga
      361 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
      421 atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
      481 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
      541 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
      601 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
      661 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
      721 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
      781 ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac aattaatcat
      841 cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc atgaccgagt
      901 acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta cgcaccctcg
      961 ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac cgccacatcg
     1021 agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg
     1081 tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag
     1141 cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg
     1201 ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt
     1261 tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg
     1321 tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacatccg
     1381 cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg
     1441 tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga cacgtgctac
     1501 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
     1561 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
     1621 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     1681 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     1741 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcattaccaa
     1801 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     1861 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagcgct
     1921 gcgatgatac cgcgagaacc acgctcaccg gctccggatt tatcagcaat aaaccagcca
     1981 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     2041 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     2101 gccatcgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     2161 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     2221 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     2281 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     2341 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     2401 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     2461 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     2521 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     2581 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     2641 tgttgaatac tcatattctt cctttttcaa tattattgaa gcatttatca gggttattgt
     2701 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggtcagtgtt
     2761 acaaccaatt aaccaattct gaacattatc gcgagcccat ttatacctga atatggctca
     2821 taacacccct tgctcatgac caaaatccct taacgtgagt tacgcgcgcg tcgttccact
     2881 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     2941 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3001 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3061 ctgttcttct agtgtagccg tagttagccc accacttcaa gaactctgta gcaccgccta
     3121 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     3181 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     3241 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     3301 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     3361 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     3421 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     3481 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     3541 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     3601 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     3661 gcgagtcagt gagcgaggaa gcggaaggcg agagtaggga actgccaggc atcaaactaa
     3721 gcagaaggcc cctgacggat ggcctttttg cgtttctaca aactctttct gtgttgtaaa
     3781 acgacggcca gtcttaagct cgggccccct gggcggttct gataacgagt aatcgttaat
     3841 ccgcaaataa cgtaaaaacc cgcttcggcg ggttttttta tggggggagt ttagggaaag
     3901 agcatttgtc agaatattta agggcgcctg tcactttgct tgatatatga gaattattta
     3961 accttataaa tgagaaaaaa gcaacgcact ttaaataaga tacgttgctt tttcgattga
     4021 tgaacaccta taattaaact attcatctat tatttatgat tttttgtata tacaatattt
     4081 ctagtttgtt aaagagaatt aagaaaataa atctcgaaaa taataaaggg aaaatcagtt
     4141 tttgatatca aaattataca tgtcaacgat aatacaaaat ataatacaaa ctataagatg
     4201 ttatcagtat ttattatcat ttagaataaa ttttgtgtcg cccttaattg tgagcggata
     4261 acaattacga gcttcatgca cagtggcgtt gactcaatat tggccattag ccatattatt
     4321 cattggttat atagcataaa tcaatattgg ctattggcca ttgcatacgt tgtatctata
     4381 tcataatatg tacatttata ttggctcatg tccaatatga ccgccatgtt ggcattgatt
     4441 attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga
     4501 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
     4561 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
     4621 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
     4681 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
     4741 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
     4801 tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
     4861 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
     4921 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
     4981 gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc agatcgcctg
     5041 gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat ccagcctccg
     5101 cggccgggaa cggtgcattg gaacgcggat tccccgtgcc aagagtgacg taagtaccgc
     5161 ctatagagtc tataggccca cccccttggc ttcgttagaa cgcggctaca attaatacat
     5221 aaccttatgt atcatacaca tacgatttag gtgacactat agaataacat ccactttgcc
     5281 tttctctcca caggtgtcca ctcccaggtc caactgcacc tcggttctat cgaaaacgcg
     5341 cctgctagcg ccaccatgaa gaccatcatt gccctgagct acatcttctg cctggtgttc
     5401 gctgcggccg ccagtcttgg aagcagagct tctctgtctg cccaagagcc tgctcaagag
     5461 gaactggtgg ccgaggaaga tcaggaccct agcgagctga atccccagac cgaagaatct
     5521 caggaccccg ctccattcct gaacagactc gtgcggccta gaagaagcgc ccctaaggga
     5581 agaaagaccc gggccagaag ggccattgcc gctcactatg aagtgcaccc tagacctgga
     5641 caggatggtg ctcaagctgg cgtggacgga acagtgtctg gatgggaaga agcccggatc
     5701 aacagcagca gccctctgcg gtataaccgg cagatcggcg agtttatcgt gaccagagcc
     5761 ggcctgtact acctgtactg ccaggtgcac tttgacgagg gcaaagccgt gtacctgaag
     5821 ctggatctgc tggtggatgg cgtgctggcc ctgagatgtc tggaagagtt ttctgctacc
     5881 gccgccagct ctctgggacc acaactgaga ctgtgtcagg tgtcaggact gctggctctg
     5941 aggcctggct ctagcctgag aatcagaaca ctgccctggg ctcacctgaa ggctgcccct
     6001 tttctgacct acttcggcct gttccaagtg catggcgcgc cggaagatca agttgatcca
     6061 agattgattg atggcaaggg cggatctctg gaggtgctgt tccagggacc aggcggaagc
     6121 ggcaatcccg gcggcgatct ggccccccag atgctgcgcg agctgcagga gaccaatgcc
     6181 gccctgcagg acgtgcgcga gctgctgcgc caccgcgtga aggagatcac cttcctgaag
     6241 aacaccgtga tggagtgcga cgcctgcggt atgcagccct ctagaggctc cggctccatc
     6301 atcccagttg aggaggagaa cccggacttc tggaaccgcg aggcagccga ggccctgggt
     6361 gccgccaaga agctgcagcc tgcacagaca gccgccaaga acctcatcat cttcctgggc
     6421 gatgggatgg gggtgtctac ggtgacagct gccaggatcc taaaagggca gaagaaggac
     6481 aaactggggc ctgagatacc cctggccatg gaccgcttcc catatgtggc tctgtccaag
     6541 acatacaatg tagacaaaca tgtgccagac agtggagcca cagccacggc ctacctgtgc
     6601 ggggtcaagg gcaacttcca gaccattggc ttgagtgcag ccgcccgctt taaccagtgc
     6661 aacacgacac gcggcaacga ggtcatctcc gtgatgaatc gggccaagaa agcagggaag
     6721 tcagtgggag tggtaaccac cacacgagtg cagcacgcct cgccagccgg cacctacgcc
     6781 cacacggtga accgcaactg gtactcggac gccgacgtgc ctgcctcggc ccgccaggag
     6841 gggtgccagg acatcgctac gcagctcatc tccaacatgg acattgacgt gatcctaggt
     6901 ggaggccgaa agtacatgtt tcgcatggga accccagacc ctgagtaccc agatgactac
     6961 agccaaggtg ggaccaggct ggacgggaag aatctggtgc aggaatggct ggcgaagcgc
     7021 cagggtgccc ggtatgtgtg gaaccgcact gagctcatgc aggcttccct ggacccgtct
     7081 gtgacccatc tcatgggtct ctttgagcct ggagacatga aatacgagat ccaccgagac
     7141 tccacactgg acccctccct gatggagatg acagaggctg ccctgcgcct gctgagcagg
     7201 aacccccgcg gcttcttcct cttcgtggag ggtggtcgca tcgaccatgg tcatcatgaa
     7261 agcagggctt accgggcact gactgagacg atcatgttcg acgacgccat tgagagggcg
     7321 ggccagctca ccagcgagga ggacacgctg agcctcgtca ctgccgacca ctcccacgtc
     7381 ttctccttcg gaggctaccc cctgcgaggg agctccatct tcgggctggc ccctggcaag
     7441 gcccgggaca ggaaggccta cacggtcctc ctatacggaa acggtccagg ctatgtgctc
     7501 aaggacggcg cccggccgga tgttaccgag agcgagagcg ggagccccga gtatcggcag
     7561 cagtcagcag tgcccctgga cgaagagacc cacgcaggcg aggacgtggc ggtgttcgcg
     7621 cgcggcccgc aggcgcacct ggttcacggc gtgcaggagc agaccttcat agcgcacgtc
     7681 atggccttcg ccgcctgcct ggagccctac accgcctgcg acctggcgcc ccccgccggc
     7741 accaccgacg ccgcgcaccc gggtggcgga ggac
//
