LOCUS       sequence-493102-        8356 bp    DNA     circular SYN 18-NOV-2025
DEFINITION  Mammalian expression of secreted protein fused to 
            Fc(DAPA)-Avi-6xHis..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8356)
  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 8356)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 18, 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-SEMA3E-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8356
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(2772..2791)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(2804..2822)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2805..2822)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3032..3056)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(3053..3256)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3082..3102)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3257..3636
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    3803..3819
                     /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(4275..4292)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4275..4291)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4390..4407)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4561..5149)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4641..4660)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5235..5254)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(5260..5279)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        5321..5412
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5413..5481,5482..6273)
                     /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(5631..5650)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    6326..6447
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6337..6356)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6391..6410
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(6577..7176)
                     /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(6660..6680)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     7157..7176
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(7195..7242)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(7290..7619)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7304..7439
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(7358..7377)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     7604..7624
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    7742..7853
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     7842..7859
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(7999..8016)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(8017..8061)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
ORIGIN
        1 gtgtacacct ggggctcccg tggctggccc ttggccttgc tgattgtctt ttcgattggg
       61 gcagccaggg ccttgttgga gactttgcac ttgtattctt tgccgttcag ccagtcctgg
      121 tgcagcacgg tcagcacgga caccaccctg taggtgctgt tgtactgctc ctctctgggc
      181 ttggtcttgg cgttgtgcac ctccacgccg tccacgtacc agttgaactt cacctctggg
      241 tcctcgtggg acacggccac caccacgcag gtcacctcgg gggtcctgct gatcatcagg
      301 gtgtccttgg gcttgggggg gaacaggaac acggaaggcc ctcccagcag ttctggagct
      361 gggcaggggg ggcaggtgtg ggtcttgtcg ccgcccggcg cgccagagtc cagggtgtgt
      421 ctaggcagtc tgtagtgctc gggcttgctc cgcagctttt tctcttgagg attggcgtac
      481 ttccacttgg aggggctcat cttcagtttc ttccgcttcc ggtcggtgca ccacactttc
      541 tcgcagtact cttccactcg ctggaagttg ctgtagccga tgagctgcag gaactctttg
      601 taccaaggct tggcgccctg agagatgctg ctctgagcag gacaaggcat tctgtggtgc
      661 cggtcctctt cgtcgtcctt gttgaacatg tcctcgacct tctcttcttc caccacctcg
      721 agggtgatct tccgcactgt gtgcacgaag ctgtgttcca cggtctggca aaagtaggtg
      781 ccggcgtcag acttgtgcag tctcaggaac agcaggccca gatccatctt gaccacgcgg
      841 tcgtctgtct tgacttcctc tttccgtgtc tcgcggccct tctgcacgaa ccaaatcact
      901 ttggcctgca gactccgagg ggtgcattcc agcagtgtgc tgttgttctc gatgccgtag
      961 gccaggtgtt cctctgtctt gtccagggcg tcgcccacaa actgctgtcc aaagcactgc
     1021 tgagcggcat ttccgtgccg cacatcctgc cgtctgaatc ttctcttggc gtgggtgcct
     1081 gtaggatagt accggctgca gctgatgcca tcccaggcac aatatggatc tctggccagg
     1141 cagcaatcgg cgcaggcgct gccatacata tcacagtggt ggaaccgcac ttgggccaca
     1201 gcagatgcag agccgatgta cagctgctgc cgcttgctgc tgatttccat gctgatgata
     1261 ggcacggggt ccttgaagat ctggagttct tccaggatca cttcttccat ggattccatc
     1321 tcttgattgt agatggtgat cactttcagc acgatgccgt tgtcggtgcc gatgaacagc
     1381 acgtcgtact ggccatcctc agcttccact ctgtccacgg cgatctgctt caggttgtac
     1441 ttgccgtcgg tcttaaccag gatgggcttc ttgtgagcgg gcttgatggc ctggtacatc
     1501 agagggtgag atctggcgaa tctgatggcg tcgtcagggt agtccttggt ggtgccgtat
     1561 ctgccgccat tcactttaga ggcgcagctg ccaggtctag gatagggcac tttgccctcg
     1621 tacacgctcc aatgatactc ggggccctct ttgtgagcat aggggccgtt gaaggcggct
     1681 ctgatgctag acatgtggta cacgcaaatg gcgtggcctc tgaagatgtt gctggtggtg
     1741 ttgaacaggc cgaagatcac agggttcttg tggtccctgg taggcagcag gaacacatct
     1801 tccagctcgt cgaagtaggt gtcgatgcca ttcatgccgg gcacgctaca cacgagtctg
     1861 gccttcagga aggtggacca cttgttgacc aggattctct ggccgccaac gtcgttcaca
     1921 cacagtctcc ccactctggt gtagatggcg tgggcattgt tctcggcttc cagggccttc
     1981 tcggtgaaga agaagtacac cttgttgtcg tccctgtcct cgttgtcggg gatcatgtag
     2041 ctgcccacga acttgggttc tttcagcagt ctctcgtcat cgtgctcggt tctgatgtgg
     2101 gccagtctgc ccatagaccg gaagatggcg gcatctctgc tccaatagtc gctgtacagg
     2161 ccggcgaaca gctcagatcc gatcagggtg ctgatgaagc tgctgctggg atcaaatgga
     2221 catctgcctc tgccgcgctc gcttctgggg ctttccaggt gaaacagagg atcttccagg
     2281 tggtagccca ctctgataaa ggcgcacaca ggatcaaaag cgccggtgcc gcaggtcagc
     2341 agatgggttc tgttgtagtg gtgcagcacc cgcacataat tggcacactc gccggcgtct
     2401 ttgcccttca tgatgcactc ttccatcttc agggctgtgc taggccagtg gatctctttg
     2461 tagccgtcgc tgatccgttc caggctcagg ctgtacacca gatctctgcc gccgacgaac
     2521 agtctctctt ggtactcgtc cagcagcatt gtgtgcaggt ccagaaagcc gaaggggctg
     2581 tgaaagatgc tggtccggtt caggttcagc agctctttgt ggctcagccg cagtctagga
     2641 tgtgttgtat cagcggcggc cgcagcgaac accaggcaga agatgtagct cagggcaatg
     2701 atggtcttca tggtggcgct agcaggcgcg ttttcgatag aaccgaggtg cagttggacc
     2761 tgggagtgga cacctgtgga gagaaaggca aagtggatgt tattctatag tgtcacctaa
     2821 atcgtatgtg tatgatacat aaggttatgt attaattgta gccgcgttct aacgaagcca
     2881 agggggtggg cctatagact ctataggcgg tacttacgtc actcttggca cggggaatcc
     2941 gcgttccaat gcaccgttcc cggccgcgga ggctggatcg gtcccggtgt cttctatgga
     3001 ggtcaaaaca gcgtggatgg cgtctccagg cgatctgacg gttcactaaa cgagctctgc
     3061 ttatatagac ctcccaccgt acacgcctac cgcccatttg cgtcaatggg gcggagttgt
     3121 tacgacattt tggaaagtcc cgttgatttt ggtgccaaaa caaactccca ttgacgtcaa
     3181 tggggtggag acttggaaat ccccgtgagt caaaccgcta tccacgccca ttgatgtact
     3241 gccaaaaccg catcaccatg gtaatagcga tgactaatac gtagatgtac tgccaagtag
     3301 gaaagtccca taaggtcatg tactgggcat aatgccaggc gggccattta ccgtcattga
     3361 cgtcaatagg gggcgtactt ggcatatgat acacttgatg tactgccaag tgggcagttt
     3421 accgtaaata ctccacccat tgacgtcaat ggaaagtccc tattggcgtt actatgggaa
     3481 catacgtcat tattgacgtc aatgggcggg ggtcgttggg cggtcagcca ggcgggccat
     3541 ttaccgtaag ttatgtaacg cggaactcca tatatgggct atgaactaat gaccccgtaa
     3601 ttgattacta ttaataacta gtcaataatc aatgccaaca tggcggtcat attggacatg
     3661 agccaatata aatgtacata ttatgatata gatacaacgt atgcaatggc caatagccaa
     3721 tattgattta tgctatataa ccaatgaata atatggctaa tggccaatat tgagtcaacg
     3781 ccactgtgca tgaagctcgt aattgttatc cgctcacaat taagggcgac acaaaattta
     3841 ttctaaatga taataaatac tgataacatc ttatagtttg tattatattt tgtattatcg
     3901 ttgacatgta taattttgat atcaaaaact gattttccct ttattatttt cgagatttat
     3961 tttcttaatt ctctttaaca aactagaaat attgtatata caaaaaatca taaataatag
     4021 atgaatagtt taattatagg tgttcatcaa tcgaaaaagc aacgtatctt atttaaagtg
     4081 cgttgctttt ttctcattta taaggttaaa taattctcat atatcaagca aagtgacagg
     4141 cgcccttaaa tattctgaca aatgctcttt ccctaaactc cccccataaa aaaacccgcc
     4201 gaagcgggtt tttacgttat ttgcggatta acgattactc gttatcagaa ccgcccaggg
     4261 ggcccgagct taagactggc cgtcgtttta caacacagaa agagtttgta gaaacgcaaa
     4321 aaggccatcc gtcaggggcc ttctgcttag tttgatgcct ggcagttccc tactctcgcc
     4381 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     4441 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     4501 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     4561 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     4621 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     4681 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     4741 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     4801 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     4861 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     4921 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtgggc
     4981 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     5041 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     5101 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     5161 gatcttttct acggggtctg acgctcagtg gaacgacgcg cgcgtaactc acgttaaggg
     5221 attttggtca tgagcaaggg gtgttatgag ccatattcag gtataaatgg gctcgcgata
     5281 atgttcagaa ttggttaatt ggttgtaaca ctgaccccta tttgtttatt tttctaaata
     5341 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     5401 aaaaggaaga atatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     5461 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     5521 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     5581 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     5641 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     5701 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     5761 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     5821 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     5881 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     5941 gacaccacga tgcctgtagc gatggcaaca acgttgcgca aactattaac tggcgaacta
     6001 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6061 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc cggagccggt
     6121 gagcgtggtt ctcgcggtat catcgcagcg ctggggccag atggtaagcc ctcccgtatc
     6181 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6241 gagataggtg cctcactgat taagcattgg taatgaccat gattacgcca agctctagct
     6301 agaggtcgac ggtatacaga catgataaga tacattgatg agtttggaca aaccacaact
     6361 agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta
     6421 accattataa gctgcaataa acaagttggg gtgggcgaag aactccagca tgagatcccc
     6481 gcgctggagg atcatccagc cggcgtcccg gaaaacgatt ccgaagccca acctttcata
     6541 gaaggcggcg gtggaatcga aatctcgtag cacgtgtcag gcaccgggct tgcgggtcat
     6601 gcaccaggtg cgcggtcctt cgggcacctc gacgtcggcg gtgacggtga agccgagccg
     6661 ctcgtagaag gggaggttgc ggggcgcgga tgtctccagg aaggcgggca ccccggcgcg
     6721 ctcggccgcc tccactccgg ggagcacgac ggcgctgccc agacccttgc cctggtggtc
     6781 gggcgagacg ccgacggtgg ccaggaacca cgcgggctcc ttgggccggt gcggcgccag
     6841 gaggccttcc atctgttgct gcgcggccag ccgggaaccg ctcaactcgg ccatgcgcgg
     6901 gccgatctcg gcgaacaccg cccccgcttc gacgctctcc ggcgtggtcc agaccgccac
     6961 cgcggcgccg tcgtccgcga cccacacctt gccgatgtcg agcccgacgc gcgtgaggaa
     7021 gagttcttgc agctcggtga cccgctcgat gtggcggtcc ggatcgacgg tgtggcgcgt
     7081 ggcggggtag tcggcgaacg cggcggcgag ggtgcgtacg gccctgggga cgtcgtcgcg
     7141 ggtggcgagg cgcaccgtgg gcttgtactc ggtcatggtt tagttcctca ccttgtcgta
     7201 ttatactatg ccgatatact atgccgatga ttaattgtca acacgtgctg atcagatccg
     7261 aaaatggata tacaagctcc cgggagcttt ttgcaaaagc ctaggcctcc aaaaaagcct
     7321 cctcactact tctggaatag ctcagaggca gaggcggcct cggcctctgc ataaataaaa
     7381 aaaattagtc agccatgggg cggagaatgg gcggaactgg gcggagttag gggcgggatg
     7441 ggcggagtta ggggcgggac tatggttgct gactaattga gatgcatgct ttgcatactt
     7501 ctgcctgctg gggagcctgg ggactttcca cacctggttg ctgactaatt gagatgcatg
     7561 ctttgcatac ttctgcctgc tggggagcct ggggactttc cacaccctaa ctgacacaca
     7621 ttccacagaa ttaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     7681 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
     7741 tgatgcaatt tcctcatttt attaggaaag gacagtggga gtggcacctt ccagggtcaa
     7801 ggaaggcacg ggggaggggc aaacaacaga tggctggcaa ctagaaggca cagtcgaggc
     7861 tgattttctc tatctattta ttttactact caagactatc ttgcaaagta gaagaaccct
     7921 gtctgcagta tgacgaccgt gtatacccga ttggcttacc gggtaacgcg ttgtgccagt
     7981 gactacaggc cggccttagt ggtgatggtg gtggtgttca tgccactcaa tcttctgagc
     8041 ctcgaaaata tcattaagac cgctgccgcc ggggctcagg ctcagggact tctgggtgta
     8101 gtggttgtgc agggcctcgt gcatcacgct gcagctgaac acgttgccct gctgccacct
     8161 ggacttgtcc acggtcagct tgctgtacag gaagaagctg ccgtcgctgt ccagcactgg
     8221 gggggtggtc ttgtagttgt tctcgggctg gccgttgctc tcccactcca cggcgatatc
     8281 gctggggtag aagcccttca ccagacaggt cagggacacc tggttcttgg tcatctcctc
     8341 ccggctgggg ggcagg
//
