LOCUS       sequence-302635-        7069 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 7069)
  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 7069)
  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-HHLA2-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..7069
                     /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             4411..4455
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4456..4473
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4613..4630)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4619..4730
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4848..4868)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4853..5182
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5033..5168
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5095..5114
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5230..5277
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5296..5895
                     /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(5296..5315)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5792..5812
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6025..6146
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6062..6081)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6116..6135
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6199..6990,6991..7059))
                     /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     6822..6841
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7060..7069,1..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 cggccgcctt tccactggcc
     2761 tttttcatct acgtgcccat gaacgagcag atcgtgatcg gcagactgga cgaggacatc
     2821 atcctgcctt ctagctttga gaggggcagc gaggtggtca tccactggaa gtaccaggac
     2881 agctacaagg tgcacagcta ctacaagggc agcgaccacc tggaatctca ggaccccaga
     2941 tacgccaacc ggaccagcct gttctacaac gagatccaga acggcaacgc ctctctgttc
     3001 ttcagaagag tgtccctgct ggatgagggc atctacacct gttacgtggg caccgccatc
     3061 caagtgatca ccaacaaggt ggtgctgaaa gtgggcgtgt tcctgacacc tgtgatgaag
     3121 tacgagaagc ggaataccaa cagcttcctg atctgcagcg tgctgagcgt gtaccccaga
     3181 cctatcatca cctggaagat ggacaacacc cctatcagcg agaacaacat ggaagagaca
     3241 ggcagcctgg acagcttcag catcaacagc cctctgaaca tcaccggcag caacagcagc
     3301 tacgagtgca ccatcgagaa cagcctgctg aagcagacat ggaccggcag atggaccatg
     3361 aaggacggcc tgcacaagat gcagtctgag cacgtgtccc tgagctgcca gcctgtgaac
     3421 gactacttca gccccaacca ggacttcaaa gtgacctggt ccagaatgaa gtccggcacc
     3481 tttagcgtgc tggcctacta cctgagcagc agccagaaca ccatcatcaa cgagagccgg
     3541 ttcagctgga acaaagagct gatcaaccag agcgacttca gcatgaacct gatggacctg
     3601 aacctgagcg acagcggcga gtacctgtgc aatatcagct ccgacgagta caccctgctg
     3661 accatccaca cagtgcacgt ggaacctagc caagaaaccg ccagccacaa tggcgcgccg
     3721 ggcggcgaca agacccacac ctgccccccc tgcccagctc cagaactgct gggagggcct
     3781 tccgtgttcc tgttcccccc caagcccaag gacaccctga tgatcagcag gacccccgag
     3841 gtgacctgcg tggtggtggc cgtgtcccac gaggacccag aggtgaagtt caactggtac
     3901 gtggacggcg tggaggtgca caacgccaag accaagccca gagaggagca gtacaacagc
     3961 acctacaggg tggtgtccgt gctgaccgtg ctgcaccagg actggctgaa cggcaaagaa
     4021 tacaagtgca aagtctccaa caaggccctg gctgccccaa tcgaaaagac aatcagcaag
     4081 gccaagggcc agccacggga gccccaggtg tacaccctgc cccccagccg ggaggagatg
     4141 accaagaacc aggtgtccct gacctgtctg gtgaagggct tctaccccag cgatatcgcc
     4201 gtggagtggg agagcaacgg ccagcccgag aacaactaca agaccacccc cccagtgctg
     4261 gacagcgacg gcagcttctt cctgtacagc aagctgaccg tggacaagtc caggtggcag
     4321 cagggcaacg tgttcagctg cagcgtgatg cacgaggccc tgcacaacca ctacacccag
     4381 aagtccctga gcctgagccc cggcggcagc ggtcttaatg atattttcga ggctcagaag
     4441 attgagtggc atgaacacca ccaccatcac cactaaggcc ggcctgtagt cactggcaca
     4501 acgcgttacc cggtaagcca atcgggtata cacggtcgtc atactgcaga cagggttctt
     4561 ctactttgca agatagtctt gagtagtaaa ataaatagat agagaaaatc agcctcgact
     4621 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     4681 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc acaacactca
     4741 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     4801 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     4861 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4921 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     4981 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     5041 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     5101 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     5161 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     5221 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     5281 gtgaggaact aaaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     5341 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     5401 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     5461 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     5521 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     5581 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     5641 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     5701 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     5761 gtgcccgcct tcctggagac atccgcgccc cgcaacctcc ccttctacga gcggctcggc
     5821 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     5881 aagcccggtg cctgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg
     5941 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     6001 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     6061 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     6121 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     6181 ttggcgtaat catggtcatt accaatgctt aatcagtgag gcacctatct cagcgatctg
     6241 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     6301 gggcttacca tctggcccca gcgctgcgat gataccgcga gaaccacgct caccggctcc
     6361 ggatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     6421 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     6481 agttaatagt ttgcgcaacg ttgttgccat cgctacaggc atcgtggtgt cacgctcgtc
     6541 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     6601 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     6661 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     6721 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     6781 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     6841 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     6901 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     6961 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     7021 aaagggaata agggcgacac ggaaatgttg aatactcata ttcttcctt
//
