LOCUS       sequence-303127-        8956 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein partial 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 8956)
  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 8956)
  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-SDK1(Trunc2)-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8956
                     /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        1767..2146
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2147..2350
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2301..2321
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2347..2371
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2581..2599
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2581..2598
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2612..2631
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             6298..6342
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6343..6360
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6500..6517)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6506..6617
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6735..6755)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6740..7069
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6920..7055
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6982..7001
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7117..7164
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7183..7782
                     /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(7183..7202)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7679..7699
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7912..8033
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7949..7968)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8003..8022
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8086..8877,8878..8946))
                     /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     8709..8728
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8947..8956,1..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 tctatatcat aatatgtaca tttatattgg
     1741 ctcatgtcca atatgaccgc catgttggca ttgattattg actagttatt aatagtaatc
     1801 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     1861 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     1921 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     1981 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     2041 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     2101 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     2161 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     2221 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     2281 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     2341 aagcagagct cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga
     2401 cctccataga agacaccggg accgatccag cctccgcggc cgggaacggt gcattggaac
     2461 gcggattccc cgtgccaaga gtgacgtaag taccgcctat agagtctata ggcccacccc
     2521 cttggcttcg ttagaacgcg gctacaatta atacataacc ttatgtatca tacacatacg
     2581 atttaggtga cactatagaa taacatccac tttgcctttc tctccacagg tgtccactcc
     2641 caggtccaac tgcacctcgg ttctatcgaa aacgcgcctg ctagcgccac catgaagacc
     2701 atcattgccc tgagctacat cttctgcctg gtgttcgctg cggccgccgg acctggtaga
     2761 cctagaggat ctcctccagg cagagccaga ccttctttgg ctcctagacc tggacctgag
     2821 cctagcagac caagagccgc tcctgaaaca tctggcggag atacagctgg cgctggaaga
     2881 tgtggtggaa gaagggccgc taaactcggc cctggtagaa gaggatggtg ggctctgctt
     2941 gccctgcagc ttcatctgct gagagccctg gctcaggatg acgtggcccc ttactttaag
     3001 acagagcccg gactgcccca gatccacctg gaaggcaata gactggtgct gacctgtctg
     3061 gccgaaggct cttggcctct ggaattcaag tggatgaggg acgacagcga gctgaccacc
     3121 tacagcagcg agtacaagta catcatcccc agcctgcaga agctggacgc cggcttttac
     3181 agatgcgtcg tgcggaatag aatgggcgcc ctgctgcaga gaaagtctga ggtgcaggtc
     3241 gcctacatgg gcagctttat ggacaccgac cagagaaaga ccgtgtctca gggcagagcc
     3301 gccatcctga atctgctgcc tatcacaagc taccccagac ctcaagtgac ctggttcaga
     3361 gagggccaca agatcatccc tagcaaccgg atcgccatca cactggaaaa ccagctcgtg
     3421 atcctggcca ccacaacatc tgatgctggc gcctattacg tgcaggccgt gaacgagaag
     3481 aacggcgaga acaagacaag ccccttcatc cacctgtcta tcgccagaga tgtgggcacc
     3541 cctgaaacaa tggcccctac aattgtggtg cctcctggca atagaagcgt ggtggccgga
     3601 agctctgaga caaccctgga atgtatcgcc agcgctagac ccgtggaaga tctgagcgtg
     3661 acctggaaga gaaacggcgt gcggatcaca tctggcctgc acagttttgg cagacggctg
     3721 accatcagca accccacaag tgccgataca ggcccttacg tgtgtgaagc tgctctgcca
     3781 ggatctgcct ttgagcctgc cagagccacc gcctttctgt tcatcatcga gcctccatac
     3841 ttcaccgccg agcctgagag cagaatcagc gccgaagtgg aagagacagt ggatatcggc
     3901 tgtcaggcca tgggagtgcc tctgcctact ctgcagtggt acaaggacgc catcagcatc
     3961 agcagactgc agaaccccag atacaaggtg ctggcttctg gcggcctgag aatccagaag
     4021 ctgaggcctg aggacagcgg catctttcag tgcttcgcct ctaacgaagg cggcgagatc
     4081 cagacacaca cctacctgga cgtgaccaat atcgcccctg tgttcaccca gaggcctgtg
     4141 gatacaaccg tgacagatgg catgaccgcc attctgagat gcgaagtgtc tggcgcccct
     4201 aagcctgcca tcacttggaa gagagagaac cacattctgg ccagcggctc cgtgcgcatc
     4261 cctagattca tgctgctgga aagcggcgga ctgcagattg ccccagtgtt tatccaggat
     4321 gccggcaact acacctgtta cgccgccaat accgagggca gcctgaatgc ctctgcaacc
     4381 ctgacagtgt ggaaccggac atccattgtg caccctccag aggaccacgt cgtgatcaag
     4441 ggaacaaccg ctacactgca ctgcggcgcc acacacgatc ctagagtgtc tctgagatac
     4501 gtgtggaaga aagacaacgt ggccctgaca cctagcagca ccagcagaat cgtggtggaa
     4561 aaagacggca gcctgctgat ctctcagact tggagcggcg acatcggcga ctactcttgc
     4621 gagattgtgt ctgaaggcgg caacgacagc cggatggcca gactggaagt gatcgaactg
     4681 cctcacagcc ctcagaacct gctggtgtct cctaatagca gccactctca cgccgtggtg
     4741 ctgagctggg ttcgaccttt tgacggcaac agccccatcc tgtactacat cgtggaactg
     4801 tccgagaaca acagcccttg gaaagtgcac ctgagcaacg tgggacctga gatgacaggc
     4861 gtgacagtgt ctggactgac ccctgccaga acctaccagt tcagagtgtg cgccgtgaat
     4921 gaagtcggca gaggccagta ctctgccgag acaagtagac tgatgctgcc cgaggaacct
     4981 cctagcgctc ctcctaagaa tatcgtggcc tccggcagga ccaaccagag cattatggtt
     5041 cagtggcagc ctccacctga gacagagcac aatggcgtgc tgagaggcta catcctgagg
     5101 tacagactgg ctggactgcc tggcgagtac cagcagagaa acatcacaag ccccgaagtg
     5161 aactactgcc tggtcaccga cctgatcatc tggacccagt atgagatcca ggtggccgcc
     5221 tataatggcg ccggactggg agtgtttagc agagccgtga ccgagtatac actgcagggc
     5281 gttccaaccg ctcctccaca gaatgtgcag accgaagccg tgaacagcac caccatccag
     5341 tttctgtgga accctccacc tcagcagttc atcaacggca tcaaccaggg ctataagctg
     5401 ctggcttggc cagccgatgc tcctgaagct gtgacagtgg tcacaatcgc cccagacttt
     5461 catggcgtgc accacggaca catcaccaac ctgaagaagt tcaccgccta cttcaccagc
     5521 gtgctgtgct ttacaacacc tggcgacggc cctccatcta cacctcagct tgtgtggacc
     5581 caagaggaca agcccggtgg cgcgccgggc ggcgacaaga cccacacctg ccccccctgc
     5641 ccagctccag aactgctggg agggccttcc gtgttcctgt tcccccccaa gcccaaggac
     5701 accctgatga tcagcaggac ccccgaggtg acctgcgtgg tggtggccgt gtcccacgag
     5761 gacccagagg tgaagttcaa ctggtacgtg gacggcgtgg aggtgcacaa cgccaagacc
     5821 aagcccagag aggagcagta caacagcacc tacagggtgg tgtccgtgct gaccgtgctg
     5881 caccaggact ggctgaacgg caaagaatac aagtgcaaag tctccaacaa ggccctggct
     5941 gccccaatcg aaaagacaat cagcaaggcc aagggccagc cacgggagcc ccaggtgtac
     6001 accctgcccc ccagccggga ggagatgacc aagaaccagg tgtccctgac ctgtctggtg
     6061 aagggcttct accccagcga tatcgccgtg gagtgggaga gcaacggcca gcccgagaac
     6121 aactacaaga ccaccccccc agtgctggac agcgacggca gcttcttcct gtacagcaag
     6181 ctgaccgtgg acaagtccag gtggcagcag ggcaacgtgt tcagctgcag cgtgatgcac
     6241 gaggccctgc acaaccacta cacccagaag tccctgagcc tgagccccgg cggcagcggt
     6301 cttaatgata ttttcgaggc tcagaagatt gagtggcatg aacaccacca ccatcaccac
     6361 taaggccggc ctgtagtcac tggcacaacg cgttacccgg taagccaatc gggtatacac
     6421 ggtcgtcata ctgcagacag ggttcttcta ctttgcaaga tagtcttgag tagtaaaata
     6481 aatagataga gaaaatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     6541 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     6601 aatgaggaaa ttgcatcaca acactcaacc ctatctcggt ctattctttt gatttataag
     6661 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     6721 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     6781 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     6841 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     6901 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     6961 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     7021 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     7081 agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag
     7141 tatatcggca tagtataata cgacaaggtg aggaactaaa ccatgaccga gtacaagccc
     7201 acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg
     7261 ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc
     7321 accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc
     7381 gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg
     7441 gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag
     7501 caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc
     7561 accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc
     7621 ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacatc cgcgccccgc
     7681 aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa
     7741 ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacacgtgct acgagatttc
     7801 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     7861 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt
     7921 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     7981 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     8041 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcattacc aatgcttaat
     8101 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     8161 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagcg ctgcgatgat
     8221 accgcgagaa ccacgctcac cggctccgga tttatcagca ataaaccagc cagccggaag
     8281 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     8341 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccatcgc
     8401 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     8461 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     8521 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     8581 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     8641 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     8701 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     8761 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     8821 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     8881 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     8941 actcatattc ttcctt
//
