LOCUS       sequence-368817-        6463 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 6463)
  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 6463)
  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-VSTM5-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6463
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     160..180
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    298..409
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     398..415
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(555..572)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(573..617)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(1791..1810)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(1823..1841)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1824..1841)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2051..2075)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2072..2275)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2101..2121)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2276..2655
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    2822..2838
                     /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(3294..3311)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3294..3310)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3409..3426)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3580..4168)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3660..3679)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4254..4273)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4279..4298)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4340..4431
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4432..4500,4501..5292)
                     /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(4650..4669)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    5345..5466
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5356..5375)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5410..5429
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5596..6195)
                     /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(5679..5699)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6176..6195
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6214..6261)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(join(6309..6463,1..175))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6323..6458
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6377..6396)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 gagttagggg cgggactatg gttgctgact aattgagatg catgctttgc atacttctgc
       61 ctgctgggga gcctggggac tttccacacc tggttgctga ctaattgaga tgcatgcttt
      121 gcatacttct gcctgctggg gagcctgggg actttccaca ccctaactga cacacattcc
      181 acagaattaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
      241 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgtgat
      301 gcaatttcct cattttatta ggaaaggaca gtgggagtgg caccttccag ggtcaaggaa
      361 ggcacggggg aggggcaaac aacagatggc tggcaactag aaggcacagt cgaggctgat
      421 tttctctatc tatttatttt actactcaag actatcttgc aaagtagaag aaccctgtct
      481 gcagtatgac gaccgtgtat acccgattgg cttaccgggt aacgcgttgt gccagtgact
      541 acaggccggc cttagtggtg atggtggtgg tgttcatgcc actcaatctt ctgagcctcg
      601 aaaatatcat taagaccgct gccgccgggg ctcaggctca gggacttctg ggtgtagtgg
      661 ttgtgcaggg cctcgtgcat cacgctgcag ctgaacacgt tgccctgctg ccacctggac
      721 ttgtccacgg tcagcttgct gtacaggaag aagctgccgt cgctgtccag cactgggggg
      781 gtggtcttgt agttgttctc gggctggccg ttgctctccc actccacggc gatatcgctg
      841 gggtagaagc ccttcaccag acaggtcagg gacacctggt tcttggtcat ctcctcccgg
      901 ctggggggca gggtgtacac ctggggctcc cgtggctggc ccttggcctt gctgattgtc
      961 ttttcgattg gggcagccag ggccttgttg gagactttgc acttgtattc tttgccgttc
     1021 agccagtcct ggtgcagcac ggtcagcacg gacaccaccc tgtaggtgct gttgtactgc
     1081 tcctctctgg gcttggtctt ggcgttgtgc acctccacgc cgtccacgta ccagttgaac
     1141 ttcacctctg ggtcctcgtg ggacacggcc accaccacgc aggtcacctc gggggtcctg
     1201 ctgatcatca gggtgtcctt gggcttgggg gggaacagga acacggaagg ccctcccagc
     1261 agttctggag ctgggcaggg ggggcaggtg tgggtcttgt cgccgcccgg cgcgccatgc
     1321 aggtcctcgt acaggatctc ggacacgtgc agcacgattg tgccaaactg gctgctgccc
     1381 agtctctctg tcacggtgat cacgtagtag ccgctgtctc tgacgcccac gctaaacagc
     1441 tggatgctgc cgttgtcgaa ggtgcacact ctgtccttgt ggctctggct gatattggcc
     1501 tgtgtgccgg gcttccattc cacgattttc tgggtgcccc agttgctgct gtaggtccac
     1561 tcgatggttg gcacgccgtg acaggagtat tccacgctca gcaggatgtc ctctttcacg
     1621 gtggcgttga tggtggcctg agggatgtac agggacacgc cttgagattg caaggcggcc
     1681 gcagcgaaca ccaggcagaa gatgtagctc agggcaatga tggtcttcat ggtggcgcta
     1741 gcaggcgcgt tttcgataga accgaggtgc agttggacct gggagtggac acctgtggag
     1801 agaaaggcaa agtggatgtt attctatagt gtcacctaaa tcgtatgtgt atgatacata
     1861 aggttatgta ttaattgtag ccgcgttcta acgaagccaa gggggtgggc ctatagactc
     1921 tataggcggt acttacgtca ctcttggcac ggggaatccg cgttccaatg caccgttccc
     1981 ggccgcggag gctggatcgg tcccggtgtc ttctatggag gtcaaaacag cgtggatggc
     2041 gtctccaggc gatctgacgg ttcactaaac gagctctgct tatatagacc tcccaccgta
     2101 cacgcctacc gcccatttgc gtcaatgggg cggagttgtt acgacatttt ggaaagtccc
     2161 gttgattttg gtgccaaaac aaactcccat tgacgtcaat ggggtggaga cttggaaatc
     2221 cccgtgagtc aaaccgctat ccacgcccat tgatgtactg ccaaaaccgc atcaccatgg
     2281 taatagcgat gactaatacg tagatgtact gccaagtagg aaagtcccat aaggtcatgt
     2341 actgggcata atgccaggcg ggccatttac cgtcattgac gtcaataggg ggcgtacttg
     2401 gcatatgata cacttgatgt actgccaagt gggcagttta ccgtaaatac tccacccatt
     2461 gacgtcaatg gaaagtccct attggcgtta ctatgggaac atacgtcatt attgacgtca
     2521 atgggcgggg gtcgttgggc ggtcagccag gcgggccatt taccgtaagt tatgtaacgc
     2581 ggaactccat atatgggcta tgaactaatg accccgtaat tgattactat taataactag
     2641 tcaataatca atgccaacat ggcggtcata ttggacatga gccaatataa atgtacatat
     2701 tatgatatag atacaacgta tgcaatggcc aatagccaat attgatttat gctatataac
     2761 caatgaataa tatggctaat ggccaatatt gagtcaacgc cactgtgcat gaagctcgta
     2821 attgttatcc gctcacaatt aagggcgaca caaaatttat tctaaatgat aataaatact
     2881 gataacatct tatagtttgt attatatttt gtattatcgt tgacatgtat aattttgata
     2941 tcaaaaactg attttccctt tattattttc gagatttatt ttcttaattc tctttaacaa
     3001 actagaaata ttgtatatac aaaaaatcat aaataataga tgaatagttt aattataggt
     3061 gttcatcaat cgaaaaagca acgtatctta tttaaagtgc gttgcttttt tctcatttat
     3121 aaggttaaat aattctcata tatcaagcaa agtgacaggc gcccttaaat attctgacaa
     3181 atgctctttc cctaaactcc ccccataaaa aaacccgccg aagcgggttt ttacgttatt
     3241 tgcggattaa cgattactcg ttatcagaac cgcccagggg gcccgagctt aagactggcc
     3301 gtcgttttac aacacagaaa gagtttgtag aaacgcaaaa aggccatccg tcaggggcct
     3361 tctgcttagt ttgatgcctg gcagttccct actctcgcct tccgcttcct cgctcactga
     3421 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     3481 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     3541 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     3601 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     3661 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     3721 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     3781 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     3841 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     3901 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     3961 gtatgtaggc ggtgctacag agttcttgaa gtggtgggct aactacggct acactagaag
     4021 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     4081 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     4141 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     4201 cgctcagtgg aacgacgcgc gcgtaactca cgttaaggga ttttggtcat gagcaagggg
     4261 tgttatgagc catattcagg tataaatggg ctcgcgataa tgttcagaat tggttaattg
     4321 gttgtaacac tgacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     4381 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagaa tatgagtatt
     4441 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     4501 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     4561 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     4621 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     4681 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     4741 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     4801 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     4861 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     4921 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagcg
     4981 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     5041 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     5101 ccggctggct ggtttattgc tgataaatcc ggagccggtg agcgtggttc tcgcggtatc
     5161 atcgcagcgc tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     5221 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     5281 aagcattggt aatgaccatg attacgccaa gctctagcta gaggtcgacg gtatacagac
     5341 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
     5401 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     5461 caagttgggg tgggcgaaga actccagcat gagatccccg cgctggagga tcatccagcc
     5521 ggcgtcccgg aaaacgattc cgaagcccaa cctttcatag aaggcggcgg tggaatcgaa
     5581 atctcgtagc acgtgtcagg caccgggctt gcgggtcatg caccaggtgc gcggtccttc
     5641 gggcacctcg acgtcggcgg tgacggtgaa gccgagccgc tcgtagaagg ggaggttgcg
     5701 gggcgcggat gtctccagga aggcgggcac cccggcgcgc tcggccgcct ccactccggg
     5761 gagcacgacg gcgctgccca gacccttgcc ctggtggtcg ggcgagacgc cgacggtggc
     5821 caggaaccac gcgggctcct tgggccggtg cggcgccagg aggccttcca tctgttgctg
     5881 cgcggccagc cgggaaccgc tcaactcggc catgcgcggg ccgatctcgg cgaacaccgc
     5941 ccccgcttcg acgctctccg gcgtggtcca gaccgccacc gcggcgccgt cgtccgcgac
     6001 ccacaccttg ccgatgtcga gcccgacgcg cgtgaggaag agttcttgca gctcggtgac
     6061 ccgctcgatg tggcggtccg gatcgacggt gtggcgcgtg gcggggtagt cggcgaacgc
     6121 ggcggcgagg gtgcgtacgg ccctggggac gtcgtcgcgg gtggcgaggc gcaccgtggg
     6181 cttgtactcg gtcatggttt agttcctcac cttgtcgtat tatactatgc cgatatacta
     6241 tgccgatgat taattgtcaa cacgtgctga tcagatccga aaatggatat acaagctccc
     6301 gggagctttt tgcaaaagcc taggcctcca aaaaagcctc ctcactactt ctggaatagc
     6361 tcagaggcag aggcggcctc ggcctctgca taaataaaaa aaattagtca gccatggggc
     6421 ggagaatggg cggaactggg cggagttagg ggcgggatgg gcg
//
