LOCUS       sequence-368738-        6619 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 6619)
  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 6619)
  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-NCR2-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6619
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     121..140
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(159..206)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(254..583)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      268..403
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(322..341)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     568..588
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    706..817
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     806..823
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(963..980)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(981..1025)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(2355..2374)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(2387..2405)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2388..2405)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2615..2639)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2636..2839)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2665..2685)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2840..3219
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    3386..3402
                     /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(3858..3875)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3858..3874)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3973..3990)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4144..4732)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4224..4243)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4818..4837)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4843..4862)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4904..4995
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4996..5064,5065..5856)
                     /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(5214..5233)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    5909..6030
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5920..5939)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5974..5993
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(6160..6619,1..140))
                     /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(6243..6263)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
ORIGIN
        1 gtgacccgct cgatgtggcg gtccggatcg acggtgtggc gcgtggcggg gtagtcggcg
       61 aacgcggcgg cgagggtgcg tacggccctg gggacgtcgt cgcgggtggc gaggcgcacc
      121 gtgggcttgt actcggtcat ggtttagttc ctcaccttgt cgtattatac tatgccgata
      181 tactatgccg atgattaatt gtcaacacgt gctgatcaga tccgaaaatg gatatacaag
      241 ctcccgggag ctttttgcaa aagcctaggc ctccaaaaaa gcctcctcac tacttctgga
      301 atagctcaga ggcagaggcg gcctcggcct ctgcataaat aaaaaaaatt agtcagccat
      361 ggggcggaga atgggcggaa ctgggcggag ttaggggcgg gatgggcgga gttaggggcg
      421 ggactatggt tgctgactaa ttgagatgca tgctttgcat acttctgcct gctggggagc
      481 ctggggactt tccacacctg gttgctgact aattgagatg catgctttgc atacttctgc
      541 ctgctgggga gcctggggac tttccacacc ctaactgaca cacattccac agaattaatt
      601 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
      661 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgtgatgc aatttcctca
      721 ttttattagg aaaggacagt gggagtggca ccttccaggg tcaaggaagg cacgggggag
      781 gggcaaacaa cagatggctg gcaactagaa ggcacagtcg aggctgattt tctctatcta
      841 tttattttac tactcaagac tatcttgcaa agtagaagaa ccctgtctgc agtatgacga
      901 ccgtgtatac ccgattggct taccgggtaa cgcgttgtgc cagtgactac aggccggcct
      961 tagtggtgat ggtggtggtg ttcatgccac tcaatcttct gagcctcgaa aatatcatta
     1021 agaccgctgc cgccggggct caggctcagg gacttctggg tgtagtggtt gtgcagggcc
     1081 tcgtgcatca cgctgcagct gaacacgttg ccctgctgcc acctggactt gtccacggtc
     1141 agcttgctgt acaggaagaa gctgccgtcg ctgtccagca ctgggggggt ggtcttgtag
     1201 ttgttctcgg gctggccgtt gctctcccac tccacggcga tatcgctggg gtagaagccc
     1261 ttcaccagac aggtcaggga cacctggttc ttggtcatct cctcccggct ggggggcagg
     1321 gtgtacacct ggggctcccg tggctggccc ttggccttgc tgattgtctt ttcgattggg
     1381 gcagccaggg ccttgttgga gactttgcac ttgtattctt tgccgttcag ccagtcctgg
     1441 tgcagcacgg tcagcacgga caccaccctg taggtgctgt tgtactgctc ctctctgggc
     1501 ttggtcttgg cgttgtgcac ctccacgccg tccacgtacc agttgaactt cacctctggg
     1561 tcctcgtggg acacggccac caccacgcag gtcacctcgg gggtcctgct gatcatcagg
     1621 gtgtccttgg gcttgggggg gaacaggaac acggaaggcc ctcccagcag ttctggagct
     1681 gggcaggggg ggcaggtgtg ggtcttgtcg ccgcccggcg cgccagcgat aggagcggca
     1741 ggtccaggtc taagtgtgct gttctgaggc tggctaggca cagggattgt agaaggagac
     1801 tcaggggcct gtctggcacc agctgttgga ggcacacagc tctgtgtctg ggtctgagag
     1861 gacaccagat ctctaggtgt ccagcttgtc tgtgtgctgg cagaggcagg agacaccacc
     1921 aggtagaacc gcacgctctt ggacacgctg ttgtcgctgg gtctgtagat ccggcaccag
     1981 tagtggccgc tatcctcttc tctcaggtcg gtcatggtca cggtaaagaa gccggcgtcg
     2041 ggatcatccc agatggtgaa tctgctggtc caggccatgg ttctgggctt gctgctggtg
     2101 accagtctga tacagacgag agcgctggcc tctttacacc agcctttctt ctcgtacagg
     2161 ctgccggtag gaggatactg gcatctgacg gtcagtgtct gtccggccac agactgcaga
     2221 acctgagctt tagactgggc ggccgcagcg aacaccaggc agaagatgta gctcagggca
     2281 atgatggtct tcatggtggc gctagcaggc gcgttttcga tagaaccgag gtgcagttgg
     2341 acctgggagt ggacacctgt ggagagaaag gcaaagtgga tgttattcta tagtgtcacc
     2401 taaatcgtat gtgtatgata cataaggtta tgtattaatt gtagccgcgt tctaacgaag
     2461 ccaagggggt gggcctatag actctatagg cggtacttac gtcactcttg gcacggggaa
     2521 tccgcgttcc aatgcaccgt tcccggccgc ggaggctgga tcggtcccgg tgtcttctat
     2581 ggaggtcaaa acagcgtgga tggcgtctcc aggcgatctg acggttcact aaacgagctc
     2641 tgcttatata gacctcccac cgtacacgcc taccgcccat ttgcgtcaat ggggcggagt
     2701 tgttacgaca ttttggaaag tcccgttgat tttggtgcca aaacaaactc ccattgacgt
     2761 caatggggtg gagacttgga aatccccgtg agtcaaaccg ctatccacgc ccattgatgt
     2821 actgccaaaa ccgcatcacc atggtaatag cgatgactaa tacgtagatg tactgccaag
     2881 taggaaagtc ccataaggtc atgtactggg cataatgcca ggcgggccat ttaccgtcat
     2941 tgacgtcaat agggggcgta cttggcatat gatacacttg atgtactgcc aagtgggcag
     3001 tttaccgtaa atactccacc cattgacgtc aatggaaagt ccctattggc gttactatgg
     3061 gaacatacgt cattattgac gtcaatgggc gggggtcgtt gggcggtcag ccaggcgggc
     3121 catttaccgt aagttatgta acgcggaact ccatatatgg gctatgaact aatgaccccg
     3181 taattgatta ctattaataa ctagtcaata atcaatgcca acatggcggt catattggac
     3241 atgagccaat ataaatgtac atattatgat atagatacaa cgtatgcaat ggccaatagc
     3301 caatattgat ttatgctata taaccaatga ataatatggc taatggccaa tattgagtca
     3361 acgccactgt gcatgaagct cgtaattgtt atccgctcac aattaagggc gacacaaaat
     3421 ttattctaaa tgataataaa tactgataac atcttatagt ttgtattata ttttgtatta
     3481 tcgttgacat gtataatttt gatatcaaaa actgattttc cctttattat tttcgagatt
     3541 tattttctta attctcttta acaaactaga aatattgtat atacaaaaaa tcataaataa
     3601 tagatgaata gtttaattat aggtgttcat caatcgaaaa agcaacgtat cttatttaaa
     3661 gtgcgttgct tttttctcat ttataaggtt aaataattct catatatcaa gcaaagtgac
     3721 aggcgccctt aaatattctg acaaatgctc tttccctaaa ctccccccat aaaaaaaccc
     3781 gccgaagcgg gtttttacgt tatttgcgga ttaacgatta ctcgttatca gaaccgccca
     3841 gggggcccga gcttaagact ggccgtcgtt ttacaacaca gaaagagttt gtagaaacgc
     3901 aaaaaggcca tccgtcaggg gccttctgct tagtttgatg cctggcagtt ccctactctc
     3961 gccttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     4021 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     4081 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4141 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4201 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4261 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4321 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4381 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4441 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4501 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4561 ggctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     4621 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     4681 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     4741 tttgatcttt tctacggggt ctgacgctca gtggaacgac gcgcgcgtaa ctcacgttaa
     4801 gggattttgg tcatgagcaa ggggtgttat gagccatatt caggtataaa tgggctcgcg
     4861 ataatgttca gaattggtta attggttgta acactgaccc ctatttgttt atttttctaa
     4921 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     4981 tgaaaaagga agaatatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     5041 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     5101 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     5161 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     5221 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     5281 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     5341 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     5401 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     5461 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     5521 cgtgacacca cgatgcctgt agcgatggca acaacgttgc gcaaactatt aactggcgaa
     5581 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     5641 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atccggagcc
     5701 ggtgagcgtg gttctcgcgg tatcatcgca gcgctggggc cagatggtaa gccctcccgt
     5761 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     5821 gctgagatag gtgcctcact gattaagcat tggtaatgac catgattacg ccaagctcta
     5881 gctagaggtc gacggtatac agacatgata agatacattg atgagtttgg acaaaccaca
     5941 actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt
     6001 gtaaccatta taagctgcaa taaacaagtt ggggtgggcg aagaactcca gcatgagatc
     6061 cccgcgctgg aggatcatcc agccggcgtc ccggaaaacg attccgaagc ccaacctttc
     6121 atagaaggcg gcggtggaat cgaaatctcg tagcacgtgt caggcaccgg gcttgcgggt
     6181 catgcaccag gtgcgcggtc cttcgggcac ctcgacgtcg gcggtgacgg tgaagccgag
     6241 ccgctcgtag aaggggaggt tgcggggcgc ggatgtctcc aggaaggcgg gcaccccggc
     6301 gcgctcggcc gcctccactc cggggagcac gacggcgctg cccagaccct tgccctggtg
     6361 gtcgggcgag acgccgacgg tggccaggaa ccacgcgggc tccttgggcc ggtgcggcgc
     6421 caggaggcct tccatctgtt gctgcgcggc cagccgggaa ccgctcaact cggccatgcg
     6481 cgggccgatc tcggcgaaca ccgcccccgc ttcgacgctc tccggcgtgg tccagaccgc
     6541 caccgcggcg ccgtcgtccg cgacccacac cttgccgatg tcgagcccga cgcgcgtgag
     6601 gaagagttct tgcagctcg
//
