LOCUS       sequence-424369-        6811 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 6811)
  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 6811)
  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-TNFRSF1B-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6811
                     /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             2387..2431
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2432..2449
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2589..2606)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2595..2706
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2824..2844)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2829..3158
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3009..3144
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3071..3090
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3206..3253
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3272..3871
                     /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(3272..3291)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3768..3788
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4001..4122
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4038..4057)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4092..4111
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4175..4966,4967..5035))
                     /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     4798..4817
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5036..5127)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5169..5188
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5194..5213
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5299..5887
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5788..5807
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6041..6058
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6156..6173
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6157..6173
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6629..6645
                     /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 ccttgcctgc tcaagtggcc tttacacctt atgctcccga
     1021 gcctggcagc acctgtagac tgagagagta ctacgaccag acagcccaga tgtgctgctc
     1081 caagtgttct cctggacagc acgccaaggt gttctgcacc aagaccagcg ataccgtgtg
     1141 cgacagctgc gaggatagca catacaccca gctgtggaat tgggtgcccg agtgtctgag
     1201 ctgtggcagc agatgtagca gcgaccaggt ggaaacccag gcctgcacca gagagcagaa
     1261 ccggatctgt acatgcagac ctggctggta ctgcgccctg tctaagcaag agggatgcag
     1321 actgtgcgcc cctctgagaa agtgtagacc cggctttgga gtggccagac caggcacaga
     1381 gacaagcgac gttgtgtgca agccttgcgc tcccggcacc tttagcaata ccaccagcag
     1441 caccgacatc tgcagacccc accagatctg caacgtggtg gccattcctg gcaatgccag
     1501 catggatgcc gtgtgcacat ctacaagccc taccagatct atggcccctg gcgctgttca
     1561 tctgcctcag cctgtgtcta ccagaagcca gcacacacag cctacacctg agcctagcac
     1621 agcccctagc acaagctttc tgctgcccat gggaccttct ccaccagccg aaggatctac
     1681 cggtgatggc gcgccgggcg gcgacaagac ccacacctgc cccccctgcc cagctccaga
     1741 actgctggga gggccttccg tgttcctgtt cccccccaag cccaaggaca ccctgatgat
     1801 cagcaggacc cccgaggtga cctgcgtggt ggtggccgtg tcccacgagg acccagaggt
     1861 gaagttcaac tggtacgtgg acggcgtgga ggtgcacaac gccaagacca agcccagaga
     1921 ggagcagtac aacagcacct acagggtggt gtccgtgctg accgtgctgc accaggactg
     1981 gctgaacggc aaagaataca agtgcaaagt ctccaacaag gccctggctg ccccaatcga
     2041 aaagacaatc agcaaggcca agggccagcc acgggagccc caggtgtaca ccctgccccc
     2101 cagccgggag gagatgacca agaaccaggt gtccctgacc tgtctggtga agggcttcta
     2161 ccccagcgat atcgccgtgg agtgggagag caacggccag cccgagaaca actacaagac
     2221 caccccccca gtgctggaca gcgacggcag cttcttcctg tacagcaagc tgaccgtgga
     2281 caagtccagg tggcagcagg gcaacgtgtt cagctgcagc gtgatgcacg aggccctgca
     2341 caaccactac acccagaagt ccctgagcct gagccccggc ggcagcggtc ttaatgatat
     2401 tttcgaggct cagaagattg agtggcatga acaccaccac catcaccact aaggccggcc
     2461 tgtagtcact ggcacaacgc gttacccggt aagccaatcg ggtatacacg gtcgtcatac
     2521 tgcagacagg gttcttctac tttgcaagat agtcttgagt agtaaaataa atagatagag
     2581 aaaatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     2641 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     2701 tgcatcacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2761 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     2821 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     2881 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     2941 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     3001 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     3061 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     3121 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     3181 ccattttcgg atctgatcag cacgtgttga caattaatca tcggcatagt atatcggcat
     3241 agtataatac gacaaggtga ggaactaaac catgaccgag tacaagccca cggtgcgcct
     3301 cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta
     3361 ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca
     3421 agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg
     3481 cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga
     3541 gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga
     3601 aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt
     3661 ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc
     3721 ggccgagcgc gccggggtgc ccgccttcct ggagacatcc gcgccccgca acctcccctt
     3781 ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac
     3841 ctggtgcatg acccgcaagc ccggtgcctg acacgtgcta cgagatttcg attccaccgc
     3901 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     3961 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     4021 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     4081 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     4141 gacctctagc tagagcttgg cgtaatcatg gtcattacca atgcttaatc agtgaggcac
     4201 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     4261 taactacgat acgggagggc ttaccatctg gccccagcgc tgcgatgata ccgcgagaac
     4321 cacgctcacc ggctccggat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     4381 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     4441 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccatcgct acaggcatcg
     4501 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     4561 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     4621 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     4681 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     4741 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     4801 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     4861 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     4921 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     4981 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatattct
     5041 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     5101 ttgaatgtat ttagaaaaat aaacaaatag gggtcagtgt tacaaccaat taaccaattc
     5161 tgaacattat cgcgagccca tttatacctg aatatggctc ataacacccc ttgctcatga
     5221 ccaaaatccc ttaacgtgag ttacgcgcgc gtcgttccac tgagcgtcag accccgtaga
     5281 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     5341 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     5401 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     5461 gtagttagcc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     5521 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     5581 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     5641 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     5701 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     5761 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     5821 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     5881 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     5941 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     6001 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     6061 agcggaaggc gagagtaggg aactgccagg catcaaacta agcagaaggc ccctgacgga
     6121 tggccttttt gcgtttctac aaactctttc tgtgttgtaa aacgacggcc agtcttaagc
     6181 tcgggccccc tgggcggttc tgataacgag taatcgttaa tccgcaaata acgtaaaaac
     6241 ccgcttcggc gggttttttt atggggggag tttagggaaa gagcatttgt cagaatattt
     6301 aagggcgcct gtcactttgc ttgatatatg agaattattt aaccttataa atgagaaaaa
     6361 agcaacgcac tttaaataag atacgttgct ttttcgattg atgaacacct ataattaaac
     6421 tattcatcta ttatttatga ttttttgtat atacaatatt tctagtttgt taaagagaat
     6481 taagaaaata aatctcgaaa ataataaagg gaaaatcagt ttttgatatc aaaattatac
     6541 atgtcaacga taatacaaaa tataatacaa actataagat gttatcagta tttattatca
     6601 tttagaataa attttgtgtc gcccttaatt gtgagcggat aacaattacg agcttcatgc
     6661 acagtggcgt tgactcaata ttggccatta gccatattat tcattggtta tatagcataa
     6721 atcaatattg gctattggcc attgcatacg ttgtatctat atcataatat gtacatttat
     6781 attggctcat gtccaatatg accgccatgt t
//
