LOCUS       sequence-303101-        8713 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 8713)
  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 8713)
  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-PXDN(Trunc3)-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8713
                     /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             6055..6099
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6100..6117
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6257..6274)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6263..6374
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6492..6512)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6497..6826
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6677..6812
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6739..6758
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6874..6921
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6940..7539
                     /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(6940..6959)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7436..7456
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7669..7790
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7706..7725)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7760..7779
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7843..8634,8635..8703))
                     /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     8466..8485
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8704..8713,1..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 cggccgccgt taatgtgcct
     2761 gacgtgtcca gaaacggcga cccttttgtg gccaccagca tcgtggaagc cattgccaca
     2821 gtggaccggg ccatcaacag caccagaaca cacctgttcg acagccggcc tagaagcccc
     2881 aatgatctgc tggccctgtt cagataccct cgggaccctt acacagtgga acaggccaga
     2941 gccggcgaga tcttcgagag aacactgcag ctgatccaag agcacgtgca gcacggcctg
     3001 atggtggatc tgaatggcac cagctaccac tacaacgacc tggtgtcccc acagtacctg
     3061 aacctgatcg ccaatctgag cggctgcaca gcccacagaa gagtgaacaa ctgcagcgac
     3121 atgtgcttcc accagaagta cagaacccac gacggcacct gtaacaacct gcagcaccct
     3181 atgtggggcg ccagcctgac agcctttgag agactgctga agtccgtgta cgagaacggc
     3241 ttcaacaccc ctcggggcat caaccctcac cggctgtata atggacacgc cctgcctatg
     3301 cctcggctgg tgtctacaac actgatcggc accgagacag tgacccctga cgagcagttt
     3361 acccacatgc tgatgcagtg gggccagttc ctggaccacg atctggattc tacagtggtg
     3421 gccctgagcc aggccagatt ttctgatggc cagcactgca gcaacgtgtg cagcaatgac
     3481 cctccttgct tcagcgtgat gatccctcct aacgacagca gagccagatc cggcgccaga
     3541 tgcatgttct ttgtgcggag cagccctgtg tgtggcagcg gcatgacaag cctgctgatg
     3601 aacagcgtgt accccagaga gcagatcaac cagctgacct cctacatcga cgcctccaac
     3661 gtgtacggca gcacagagca cgaggcccgc tctattagag atctggcctc tcacagaggc
     3721 ctgctgagac agggaatcgt gcagagaagc ggcaagcccc tgctgccttt tgctaccgga
     3781 cctccaaccg agtgcatgcg ggatgagaac gagagcccca ttccttgctt tctggccggc
     3841 gaccacagag ccaatgaaca gctgggcctg accagcatgc acaccctgtg gttcagagag
     3901 cacaaccgga tcgccaccga gctgctgaag ctgaaccctc actgggacgg cgacaccatc
     3961 tactacgaga cacggaagat cgtgggcgcc gagatccagc acatcaccta tcagcactgg
     4021 ctgcccaaga tcctgggcga agtgggaatg agaaccctgg gcgagtacca cggctacgac
     4081 cctggaatca acgccggcat cttcaacgcc tttgccaccg ccgccttcag attcggccac
     4141 acactggtca accctctgct gtacagactg gacgagaact tccagcctat cgctcaggac
     4201 catctgcctc tgcacaaggc tttcttcagc cccttccgga tcgtgaacga aggcggcatt
     4261 gacccactgc tgagaggcct gtttggagtg gccggcaaga tgagagtgcc tagccagctg
     4321 ctcaataccg agctgaccga gcggctgttt agcatggccc atacagtggc tctggacctg
     4381 gccgccatca atatccagag aggcagagat cacggcatcc ctccatacca cgactaccgg
     4441 gtgtactgca atctgtccgc cgctcacacc ttcgaggacc tgaagaacga gatcaagaac
     4501 cccgagatcc gcgagaagct gaagcggctg tatggctcta ccctgaacat cgatctgttc
     4561 cccgctctgg tggtggaaga tctggtgcct ggaagcagac tgggccccac actgatgtgt
     4621 ctgctgagca cccagttcaa gcggctgaga gatggcgacc ggctttggta cgaaaacccc
     4681 ggtgtcttta gccccgctca gctgacccag atcaagcaga ccagcctggc cagaatcctg
     4741 tgcgacaacg ccgacaacat caccagagtg cagagcgacg tgttcagagt ggccgagttt
     4801 cctcacggct atggcagctg tgacgagatc cccagagtgg acctgagagt gtggcaggac
     4861 tgctgcgagg actgcagaac cagaggccag ttcaatgcct tcagctatca cttccggggc
     4921 agaagaagcc tcgagttcag ctaccaagag gacaagccca ccaagaaaac ccggcctcgg
     4981 aagatcccaa gcgttggcag acaaggcgag cacctgagca atagcaccag cgccttctcc
     5041 accagatccg atgccagcgg caccaacgac ttccgcgagt tcgtgctgga aatgcagaaa
     5101 accatcaccg acctgcggac ccagattaag aagctggaaa gccgcctgag caccaccgag
     5161 tgtgttgatg ctggcggaga gagccacgcc aacaacacca agtggaagaa ggacgcctgc
     5221 accatctgcg agtgcaagga tggccaagtg acctgctttg tggaagcctg tcctcctgcc
     5281 acttgtgccg tgcctgtgaa tatccctggc gcctgctgtc ctgtgtgcct gcagaaaaga
     5341 gccgaggaaa agcctggcgc gccgggcggc gacaagaccc acacctgccc cccctgccca
     5401 gctccagaac tgctgggagg gccttccgtg ttcctgttcc cccccaagcc caaggacacc
     5461 ctgatgatca gcaggacccc cgaggtgacc tgcgtggtgg tggccgtgtc ccacgaggac
     5521 ccagaggtga agttcaactg gtacgtggac ggcgtggagg tgcacaacgc caagaccaag
     5581 cccagagagg agcagtacaa cagcacctac agggtggtgt ccgtgctgac cgtgctgcac
     5641 caggactggc tgaacggcaa agaatacaag tgcaaagtct ccaacaaggc cctggctgcc
     5701 ccaatcgaaa agacaatcag caaggccaag ggccagccac gggagcccca ggtgtacacc
     5761 ctgcccccca gccgggagga gatgaccaag aaccaggtgt ccctgacctg tctggtgaag
     5821 ggcttctacc ccagcgatat cgccgtggag tgggagagca acggccagcc cgagaacaac
     5881 tacaagacca cccccccagt gctggacagc gacggcagct tcttcctgta cagcaagctg
     5941 accgtggaca agtccaggtg gcagcagggc aacgtgttca gctgcagcgt gatgcacgag
     6001 gccctgcaca accactacac ccagaagtcc ctgagcctga gccccggcgg cagcggtctt
     6061 aatgatattt tcgaggctca gaagattgag tggcatgaac accaccacca tcaccactaa
     6121 ggccggcctg tagtcactgg cacaacgcgt tacccggtaa gccaatcggg tatacacggt
     6181 cgtcatactg cagacagggt tcttctactt tgcaagatag tcttgagtag taaaataaat
     6241 agatagagaa aatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     6301 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     6361 gaggaaattg catcacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     6421 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     6481 attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     6541 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     6601 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     6661 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     6721 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt
     6781 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc
     6841 ttgtatatcc attttcggat ctgatcagca cgtgttgaca attaatcatc ggcatagtat
     6901 atcggcatag tataatacga caaggtgagg aactaaacca tgaccgagta caagcccacg
     6961 gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc
     7021 gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc
     7081 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     7141 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     7201 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     7261 cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     7321 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     7381 gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacatccgc gccccgcaac
     7441 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     7501 ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac acgtgctacg agatttcgat
     7561 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     7621 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     7681 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     7741 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     7801 ataccgtcga cctctagcta gagcttggcg taatcatggt cattaccaat gcttaatcag
     7861 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     7921 cgtgtagata actacgatac gggagggctt accatctggc cccagcgctg cgatgatacc
     7981 gcgagaacca cgctcaccgg ctccggattt atcagcaata aaccagccag ccggaagggc
     8041 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     8101 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccatcgctac
     8161 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     8221 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     8281 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     8341 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     8401 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     8461 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     8521 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     8581 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     8641 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     8701 catattcttc ctt
//
