LOCUS       sequence-430445-        6724 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 6724)
  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 6724)
  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-TMEM25-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6724
                     /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             2300..2344
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2345..2362
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2502..2519)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2508..2619
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2737..2757)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2742..3071
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2922..3057
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2984..3003
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3119..3166
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3185..3784
                     /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(3185..3204)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3681..3701
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3914..4035
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3951..3970)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4005..4024
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4088..4879,4880..4948))
                     /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     4711..4730
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4949..5040)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5082..5101
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5107..5126
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5212..5800
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5701..5720
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5954..5971
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6069..6086
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6070..6086
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6542..6558
                     /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 ccgaactgga acctcagatt gatggacaga catgggccga
     1021 gagagccctg agagaaaacg agagacacgc cttcacctgt agagtggctg gtggccctgg
     1081 aacacctaga ctggcttggt atctggacgg acagctgcaa gaggccagca caagcagact
     1141 gctgtctgtt ggcggagagg cctttagcgg cggcacaagc acttttaccg tgacagccca
     1201 cagagcccag cacgagctga attgcagcct gcaagatccc agaagcggca gatcagccaa
     1261 tgccagcgtg atcctgaacg tgcagttcaa gcctgagatc gcccaagtgg gcgccaagta
     1321 ccaagaagct caaggacctg gactgctggt ggtgctgttt gccctcgtta gagccaatcc
     1381 tcctgccaac gtgacctgga tcgatcagga tggccctgtg accgtgaata ccagcgactt
     1441 tctggtgctg gacgcccaga attacccctg gctgaccaat cacacagtgc agctccagct
     1501 gagatctctg gcccacaatc tgagcgtggt ggccacaaat gatgtgggcg tgacaagcgc
     1561 ttctctgcct gctcctggac tcctggccac aagagttgaa ggcgcgccgg gcggcgacaa
     1621 gacccacacc tgccccccct gcccagctcc agaactgctg ggagggcctt ccgtgttcct
     1681 gttccccccc aagcccaagg acaccctgat gatcagcagg acccccgagg tgacctgcgt
     1741 ggtggtggcc gtgtcccacg aggacccaga ggtgaagttc aactggtacg tggacggcgt
     1801 ggaggtgcac aacgccaaga ccaagcccag agaggagcag tacaacagca cctacagggt
     1861 ggtgtccgtg ctgaccgtgc tgcaccagga ctggctgaac ggcaaagaat acaagtgcaa
     1921 agtctccaac aaggccctgg ctgccccaat cgaaaagaca atcagcaagg ccaagggcca
     1981 gccacgggag ccccaggtgt acaccctgcc ccccagccgg gaggagatga ccaagaacca
     2041 ggtgtccctg acctgtctgg tgaagggctt ctaccccagc gatatcgccg tggagtggga
     2101 gagcaacggc cagcccgaga acaactacaa gaccaccccc ccagtgctgg acagcgacgg
     2161 cagcttcttc ctgtacagca agctgaccgt ggacaagtcc aggtggcagc agggcaacgt
     2221 gttcagctgc agcgtgatgc acgaggccct gcacaaccac tacacccaga agtccctgag
     2281 cctgagcccc ggcggcagcg gtcttaatga tattttcgag gctcagaaga ttgagtggca
     2341 tgaacaccac caccatcacc actaaggccg gcctgtagtc actggcacaa cgcgttaccc
     2401 ggtaagccaa tcgggtatac acggtcgtca tactgcagac agggttcttc tactttgcaa
     2461 gatagtcttg agtagtaaaa taaatagata gagaaaatca gcctcgactg tgccttctag
     2521 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     2581 tcccactgtc ctttcctaat aaaatgagga aattgcatca caacactcaa ccctatctcg
     2641 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     2701 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     2761 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     2821 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     2881 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     2941 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     3001 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3061 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtgt
     3121 tgacaattaa tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta
     3181 aaccatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc
     3241 cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc
     3301 ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct
     3361 cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc
     3421 ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag
     3481 cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa
     3541 ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct
     3601 gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt
     3661 cctggagaca tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac
     3721 cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc
     3781 ctgacacgtg ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     3841 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     3901 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     3961 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     4021 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc
     4081 atggtcatta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     4141 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     4201 ctggccccag cgctgcgatg ataccgcgag aaccacgctc accggctccg gatttatcag
     4261 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     4321 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     4381 tgcgcaacgt tgttgccatc gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     4441 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     4501 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     4561 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     4621 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     4681 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     4741 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     4801 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     4861 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     4921 gggcgacacg gaaatgttga atactcatat tcttcctttt tcaatattat tgaagcattt
     4981 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     5041 taggggtcag tgttacaacc aattaaccaa ttctgaacat tatcgcgagc ccatttatac
     5101 ctgaatatgg ctcataacac cccttgctca tgaccaaaat cccttaacgt gagttacgcg
     5161 cgcgtcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     5221 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     5281 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     5341 gcagatacca aatactgttc ttctagtgta gccgtagtta gcccaccact tcaagaactc
     5401 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     5461 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     5521 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     5581 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     5641 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     5701 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     5761 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     5821 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     5881 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     5941 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa ggcgagagta gggaactgcc
     6001 aggcatcaaa ctaagcagaa ggcccctgac ggatggcctt tttgcgtttc tacaaactct
     6061 ttctgtgttg taaaacgacg gccagtctta agctcgggcc ccctgggcgg ttctgataac
     6121 gagtaatcgt taatccgcaa ataacgtaaa aacccgcttc ggcgggtttt tttatggggg
     6181 gagtttaggg aaagagcatt tgtcagaata tttaagggcg cctgtcactt tgcttgatat
     6241 atgagaatta tttaacctta taaatgagaa aaaagcaacg cactttaaat aagatacgtt
     6301 gctttttcga ttgatgaaca cctataatta aactattcat ctattattta tgattttttg
     6361 tatatacaat atttctagtt tgttaaagag aattaagaaa ataaatctcg aaaataataa
     6421 agggaaaatc agtttttgat atcaaaatta tacatgtcaa cgataataca aaatataata
     6481 caaactataa gatgttatca gtatttatta tcatttagaa taaattttgt gtcgccctta
     6541 attgtgagcg gataacaatt acgagcttca tgcacagtgg cgttgactca atattggcca
     6601 ttagccatat tattcattgg ttatatagca taaatcaata ttggctattg gccattgcat
     6661 acgttgtatc tatatcataa tatgtacatt tatattggct catgtccaat atgaccgcca
     6721 tgtt
//
