LOCUS       sequence-368608-        6529 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 6529)
  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 6529)
  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-TNFRSF19-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6529
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     246..266
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    384..495
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     484..501
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(641..658)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(659..703)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     RBS             1479..1487
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     primer_bind     complement(1943..1962)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(1975..1993)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1976..1993)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2203..2227)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2224..2427)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2253..2273)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2428..2807
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    2974..2990
                     /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)."
     primer_bind     complement(3446..3463)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3446..3462)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3561..3578)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3732..4320)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3812..3831)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4406..4425)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4431..4450)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4492..4583
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4584..4652,4653..5444)
                     /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     complement(4802..4821)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    5497..5618
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5508..5527)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5562..5581
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5748..6347)
                     /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(5831..5851)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6328..6347
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6366..6413)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(join(6461..6529,1..261))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6475..6529,1..81)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(join(6529,1..19))
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 ctcggcctct gcataaataa aaaaaattag tcagccatgg ggcggagaat gggcggaact
       61 gggcggagtt aggggcggga tgggcggagt taggggcggg actatggttg ctgactaatt
      121 gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc cacacctggt
      181 tgctgactaa ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt
      241 tccacaccct aactgacaca cattccacag aattaattcg cgttaaattt ttgttaaatc
      301 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
      361 accgagatag ggttgagtgt tgtgatgcaa tttcctcatt ttattaggaa aggacagtgg
      421 gagtggcacc ttccagggtc aaggaaggca cgggggaggg gcaaacaaca gatggctggc
      481 aactagaagg cacagtcgag gctgattttc tctatctatt tattttacta ctcaagacta
      541 tcttgcaaag tagaagaacc ctgtctgcag tatgacgacc gtgtataccc gattggctta
      601 ccgggtaacg cgttgtgcca gtgactacag gccggcctta gtggtgatgg tggtggtgtt
      661 catgccactc aatcttctga gcctcgaaaa tatcattaag accgctgccg ccggggctca
      721 ggctcaggga cttctgggtg tagtggttgt gcagggcctc gtgcatcacg ctgcagctga
      781 acacgttgcc ctgctgccac ctggacttgt ccacggtcag cttgctgtac aggaagaagc
      841 tgccgtcgct gtccagcact gggggggtgg tcttgtagtt gttctcgggc tggccgttgc
      901 tctcccactc cacggcgata tcgctggggt agaagccctt caccagacag gtcagggaca
      961 cctggttctt ggtcatctcc tcccggctgg ggggcagggt gtacacctgg ggctcccgtg
     1021 gctggccctt ggccttgctg attgtctttt cgattggggc agccagggcc ttgttggaga
     1081 ctttgcactt gtattctttg ccgttcagcc agtcctggtg cagcacggtc agcacggaca
     1141 ccaccctgta ggtgctgttg tactgctcct ctctgggctt ggtcttggcg ttgtgcacct
     1201 ccacgccgtc cacgtaccag ttgaacttca cctctgggtc ctcgtgggac acggccacca
     1261 ccacgcaggt cacctcgggg gtcctgctga tcatcagggt gtccttgggc ttggggggga
     1321 acaggaacac ggaaggccct cccagcagtt ctggagctgg gcaggggggg caggtgtggg
     1381 tcttgtcgcc gcccggcgcg ccaagagctg tgtctcttgg agagctggcg gtgctggcga
     1441 tcttgaccag gttcactttg ctggcgcagt gaggttcata aggaggtgga gggtcgccac
     1501 atggcacgca ttccatgtcc tggaatccca cgagcttggt ctttctgtag aagccgggca
     1561 gacaatcgcc acagatggca tcggaggtgg cgctgcaatt ggccttctgg aatctgttga
     1621 ccacggcgca gtccagacaa ggcttgcact tctggaagcc ccagtcctct ttgaatctgt
     1681 gcagccggca ggtcacacac tgggcatcct cgccgtatcc gaagccacac tctttgctca
     1741 gttccatgcc agggccacac tgattacagg gcacgcagtt gccggatctg tcccggaact
     1801 cttgctgtct gcaatcgcca gattcggcgg ccgcagcgaa caccaggcag aagatgtagc
     1861 tcagggcaat gatggtcttc atggtggcgc tagcaggcgc gttttcgata gaaccgaggt
     1921 gcagttggac ctgggagtgg acacctgtgg agagaaaggc aaagtggatg ttattctata
     1981 gtgtcaccta aatcgtatgt gtatgataca taaggttatg tattaattgt agccgcgttc
     2041 taacgaagcc aagggggtgg gcctatagac tctataggcg gtacttacgt cactcttggc
     2101 acggggaatc cgcgttccaa tgcaccgttc ccggccgcgg aggctggatc ggtcccggtg
     2161 tcttctatgg aggtcaaaac agcgtggatg gcgtctccag gcgatctgac ggttcactaa
     2221 acgagctctg cttatataga cctcccaccg tacacgccta ccgcccattt gcgtcaatgg
     2281 ggcggagttg ttacgacatt ttggaaagtc ccgttgattt tggtgccaaa acaaactccc
     2341 attgacgtca atggggtgga gacttggaaa tccccgtgag tcaaaccgct atccacgccc
     2401 attgatgtac tgccaaaacc gcatcaccat ggtaatagcg atgactaata cgtagatgta
     2461 ctgccaagta ggaaagtccc ataaggtcat gtactgggca taatgccagg cgggccattt
     2521 accgtcattg acgtcaatag ggggcgtact tggcatatga tacacttgat gtactgccaa
     2581 gtgggcagtt taccgtaaat actccaccca ttgacgtcaa tggaaagtcc ctattggcgt
     2641 tactatggga acatacgtca ttattgacgt caatgggcgg gggtcgttgg gcggtcagcc
     2701 aggcgggcca tttaccgtaa gttatgtaac gcggaactcc atatatgggc tatgaactaa
     2761 tgaccccgta attgattact attaataact agtcaataat caatgccaac atggcggtca
     2821 tattggacat gagccaatat aaatgtacat attatgatat agatacaacg tatgcaatgg
     2881 ccaatagcca atattgattt atgctatata accaatgaat aatatggcta atggccaata
     2941 ttgagtcaac gccactgtgc atgaagctcg taattgttat ccgctcacaa ttaagggcga
     3001 cacaaaattt attctaaatg ataataaata ctgataacat cttatagttt gtattatatt
     3061 ttgtattatc gttgacatgt ataattttga tatcaaaaac tgattttccc tttattattt
     3121 tcgagattta ttttcttaat tctctttaac aaactagaaa tattgtatat acaaaaaatc
     3181 ataaataata gatgaatagt ttaattatag gtgttcatca atcgaaaaag caacgtatct
     3241 tatttaaagt gcgttgcttt tttctcattt ataaggttaa ataattctca tatatcaagc
     3301 aaagtgacag gcgcccttaa atattctgac aaatgctctt tccctaaact ccccccataa
     3361 aaaaacccgc cgaagcgggt ttttacgtta tttgcggatt aacgattact cgttatcaga
     3421 accgcccagg gggcccgagc ttaagactgg ccgtcgtttt acaacacaga aagagtttgt
     3481 agaaacgcaa aaaggccatc cgtcaggggc cttctgctta gtttgatgcc tggcagttcc
     3541 ctactctcgc cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     3601 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     3661 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     3721 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     3781 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     3841 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     3901 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     3961 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4021 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4081 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4141 aagtggtggg ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     4201 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4261 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4321 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgacgc gcgcgtaact
     4381 cacgttaagg gattttggtc atgagcaagg ggtgttatga gccatattca ggtataaatg
     4441 ggctcgcgat aatgttcaga attggttaat tggttgtaac actgacccct atttgtttat
     4501 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     4561 aataatattg aaaaaggaag aatatgagta ttcaacattt ccgtgtcgcc cttattccct
     4621 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     4681 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     4741 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     4801 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     4861 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     4921 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     4981 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     5041 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     5101 acgacgagcg tgacaccacg atgcctgtag cgatggcaac aacgttgcgc aaactattaa
     5161 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     5221 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     5281 ccggagccgg tgagcgtggt tctcgcggta tcatcgcagc gctggggcca gatggtaagc
     5341 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     5401 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaatgacca tgattacgcc
     5461 aagctctagc tagaggtcga cggtatacag acatgataag atacattgat gagtttggac
     5521 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     5581 ctttatttgt aaccattata agctgcaata aacaagttgg ggtgggcgaa gaactccagc
     5641 atgagatccc cgcgctggag gatcatccag ccggcgtccc ggaaaacgat tccgaagccc
     5701 aacctttcat agaaggcggc ggtggaatcg aaatctcgta gcacgtgtca ggcaccgggc
     5761 ttgcgggtca tgcaccaggt gcgcggtcct tcgggcacct cgacgtcggc ggtgacggtg
     5821 aagccgagcc gctcgtagaa ggggaggttg cggggcgcgg atgtctccag gaaggcgggc
     5881 accccggcgc gctcggccgc ctccactccg gggagcacga cggcgctgcc cagacccttg
     5941 ccctggtggt cgggcgagac gccgacggtg gccaggaacc acgcgggctc cttgggccgg
     6001 tgcggcgcca ggaggccttc catctgttgc tgcgcggcca gccgggaacc gctcaactcg
     6061 gccatgcgcg ggccgatctc ggcgaacacc gcccccgctt cgacgctctc cggcgtggtc
     6121 cagaccgcca ccgcggcgcc gtcgtccgcg acccacacct tgccgatgtc gagcccgacg
     6181 cgcgtgagga agagttcttg cagctcggtg acccgctcga tgtggcggtc cggatcgacg
     6241 gtgtggcgcg tggcggggta gtcggcgaac gcggcggcga gggtgcgtac ggccctgggg
     6301 acgtcgtcgc gggtggcgag gcgcaccgtg ggcttgtact cggtcatggt ttagttcctc
     6361 accttgtcgt attatactat gccgatatac tatgccgatg attaattgtc aacacgtgct
     6421 gatcagatcc gaaaatggat atacaagctc ccgggagctt tttgcaaaag cctaggcctc
     6481 caaaaaagcc tcctcactac ttctggaata gctcagaggc agaggcggc
//
