LOCUS       sequence-462340-        8278 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to Fc(DAPA)-Avi-6xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8278)
  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 8278)
  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-AZGP1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8278
                     /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        3105..3484
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3485..3688
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3639..3659
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3685..3709
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        3919..3937
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3919..3936
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     3950..3969
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             5620..5664
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5665..5682
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5822..5839)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5828..5939
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6057..6077)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6062..6391
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6242..6377
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6304..6323
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6439..6486
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6505..7104
                     /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(6505..6524)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7001..7021
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7234..7355
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7271..7290)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7325..7344
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7408..8199,8200..8268))
                     /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     8031..8050
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8269..8278,1..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 ctgatgaatc ccctaatgat tttggtaaaa
     1741 atcattaagt taaggtggat acacatcttg tcatatgatc aaatggtttc gcgaaaaatc
     1801 aataatcaga caacaagatg tgcgaactcg atattttaca cgactctctt taccaattct
     1861 gccccgaatt acacttaaaa cgactcaaca gcttaacgtt ggcttgccac gcattacttg
     1921 actgtaaaac tctcactctt accgaacttg gccgtaacct gccaaccaaa gcgagaacaa
     1981 aacataacat caaacgaatc gaccgattgt taggtaatcg tcacctccac aaagagcgac
     2041 tcgctgtata ccgttggcat gctagcttta tctgttcggg caatacgatg cccattgtac
     2101 ttgttgactg gtctgatatt cgtgagcaaa aacgacttat ggtattgcga gcttcagtcg
     2161 cactacacgg tcgttctgtt actctttatg agaaagcgtt cccgctttca gagcaatgtt
     2221 caaagaaagc tcatgaccaa tttctagccg accttgcgag cattctaccg agtaacacca
     2281 caccgctcat tgtcagtgat gctggcttta aagtgccatg gtataaatcc gttgagaagc
     2341 tgggttggta ctggttaagt cgagtaagag gaaaagtaca atatgcagac ctaggagcgg
     2401 aaaactggaa acctatcagc aacttacatg atatgtcatc tagtcactca aagactttag
     2461 gctataagag gctgactaaa agcaatccaa tctcatgcca aattctattg tataaatctc
     2521 gctctaaagg ccgaaaaaat cagcgctcga cacggactca ttgtcaccac ccgtcaccta
     2581 aaatctactc agcgtcggca aaggagccat gggttctagc aactaactta cctgttgaaa
     2641 ttcgaacacc caaacaactt gttaatatct attcgaagcg aatgcagatt gaagaaacct
     2701 tccgagactt gaaaagtcct gcctacggac taggcctacg ccatagccga acgagcagct
     2761 cagagcgttt tgatatcatg ctgctaatcg ccctgatgct tcaactaaca tgttggcttg
     2821 cgggcgttca tgctcagaaa caaggttggg acaagcactt ccaggctaac acagtcagaa
     2881 atcgaaacgt actctcaaca gttcgcttag gcatggaagt tttgcggcat tctggctaca
     2941 caataacaag ggaagactta ctcgtggctg caaccctact agctcaaaat ttattcacac
     3001 atggttacgc tttggggaaa ttatgagggg atctctcagt acgttgtatc tatatcataa
     3061 tatgtacatt tatattggct catgtccaat atgaccgcca tgttggcatt gattattgac
     3121 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
     3181 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
     3241 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
     3301 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
     3361 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
     3421 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
     3481 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
     3541 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
     3601 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
     3661 acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagatcg cctggagacg
     3721 ccatccacgc tgttttgacc tccatagaag acaccgggac cgatccagcc tccgcggccg
     3781 ggaacggtgc attggaacgc ggattccccg tgccaagagt gacgtaagta ccgcctatag
     3841 agtctatagg cccaccccct tggcttcgtt agaacgcggc tacaattaat acataacctt
     3901 atgtatcata cacatacgat ttaggtgaca ctatagaata acatccactt tgcctttctc
     3961 tccacaggtg tccactccca ggtccaactg cacctcggtt ctatcgaaaa cgcgcctgct
     4021 agcgccacca tgaagaccat cattgccctg agctacatct tctgcctggt gttcgctgcg
     4081 gccgcccaag agaatcagga tggcagatac agcctgacct acatctacac cggcctgagc
     4141 aagcacgtgg aagatgtgcc tgcttttcag gccctgggca gcctgaacga cctgcagttc
     4201 ttccggtaca acagcaagga ccggaagtcc cagcctatgg gcctttggag acaggtggaa
     4261 ggcatggaag attggaagca ggacagccag ctgcagaagg cccgcgagga tatcttcatg
     4321 gaaaccctga aggacatcgt cgagtactac aacgacagca acggcagcca cgtgctgcag
     4381 ggcagatttg gatgcgagat cgagaacaac agaagcagcg gcgccttctg gaagtactac
     4441 tacgatggca aggactacat cgagttcaac aaagaaatcc ccgcctgggt gcccttcgat
     4501 cctgccgctc agattaccaa gcagaagtgg gaagccgagc ctgtgtacgt gcagagagcc
     4561 aaggcctacc tggaagagga atgccccgcc acactgcgga agtacctgaa gtacagcaag
     4621 aacatcctgg accggcagga ccctcctagc gtggtggtta catctcatca ggcccctggc
     4681 gagaagaaaa agctgaagtg cctggcctac gacttctacc ccggcaagat cgatgtgcac
     4741 tggacaagag ccggcgaggt gcaagaacct gaactgagag gggatgtgct gcacaacggc
     4801 aatggcacat accagtcttg ggtcgtcgtg gcagtgccac ctcaggatac agccccttac
     4861 tcttgtcacg tgcagcactc tagtctggcc cagcctctgg ttgtgccttg ggaagcttct
     4921 ggcgcgccgg gcggcgacaa gacccacacc tgccccccct gcccagctcc agaactgctg
     4981 ggagggcctt ccgtgttcct gttccccccc aagcccaagg acaccctgat gatcagcagg
     5041 acccccgagg tgacctgcgt ggtggtggcc gtgtcccacg aggacccaga ggtgaagttc
     5101 aactggtacg tggacggcgt ggaggtgcac aacgccaaga ccaagcccag agaggagcag
     5161 tacaacagca cctacagggt ggtgtccgtg ctgaccgtgc tgcaccagga ctggctgaac
     5221 ggcaaagaat acaagtgcaa agtctccaac aaggccctgg ctgccccaat cgaaaagaca
     5281 atcagcaagg ccaagggcca gccacgggag ccccaggtgt acaccctgcc ccccagccgg
     5341 gaggagatga ccaagaacca ggtgtccctg acctgtctgg tgaagggctt ctaccccagc
     5401 gatatcgccg tggagtggga gagcaacggc cagcccgaga acaactacaa gaccaccccc
     5461 ccagtgctgg acagcgacgg cagcttcttc ctgtacagca agctgaccgt ggacaagtcc
     5521 aggtggcagc agggcaacgt gttcagctgc agcgtgatgc acgaggccct gcacaaccac
     5581 tacacccaga agtccctgag cctgagcccc ggcggcagcg gtcttaatga tattttcgag
     5641 gctcagaaga ttgagtggca tgaacaccac caccatcacc actaaggccg gcctgtagtc
     5701 actggcacaa cgcgttaccc ggtaagccaa tcgggtatac acggtcgtca tactgcagac
     5761 agggttcttc tactttgcaa gatagtcttg agtagtaaaa taaatagata gagaaaatca
     5821 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
     5881 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatca
     5941 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
     6001 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa
     6061 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     6121 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     6181 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     6241 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     6301 ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg
     6361 aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt
     6421 cggatctgat cagcacgtgt tgacaattaa tcatcggcat agtatatcgg catagtataa
     6481 tacgacaagg tgaggaacta aaccatgacc gagtacaagc ccacggtgcg cctcgccacc
     6541 cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc
     6601 acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc
     6661 ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg
     6721 gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc
     6781 ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc
     6841 ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc
     6901 gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag
     6961 cgcgccgggg tgcccgcctt cctggagaca tccgcgcccc gcaacctccc cttctacgag
     7021 cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc
     7081 atgacccgca agcccggtgc ctgacacgtg ctacgagatt tcgattccac cgccgccttc
     7141 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     7201 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     7261 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     7321 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     7381 agctagagct tggcgtaatc atggtcatta ccaatgctta atcagtgagg cacctatctc
     7441 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     7501 gatacgggag ggcttaccat ctggccccag cgctgcgatg ataccgcgag aaccacgctc
     7561 accggctccg gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     7621 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     7681 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatc gctacaggca tcgtggtgtc
     7741 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     7801 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     7861 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     7921 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     7981 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     8041 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     8101 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     8161 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     8221 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatat tcttcctt
//
