LOCUS       sequence-424368-        6730 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 6730)
  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 6730)
  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-CD80-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6730
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        77..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        815..833
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     815..832
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     846..865
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2306..2350
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2351..2368
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2508..2525)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2514..2625
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2743..2763)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2748..3077
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2928..3063
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2990..3009
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3125..3172
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3191..3790
                     /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(3191..3210)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3687..3707
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3920..4041
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3957..3976)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4011..4030
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4094..4885,4886..4954))
                     /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     4717..4736
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4955..5046)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5088..5107
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5113..5132
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5218..5806
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5707..5726
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5960..5977
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6075..6092
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6076..6092
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6548..6564
                     /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)."
ORIGIN
        1 ggcattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat
      661 ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc aagagtgacg
      721 taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa cgcggctaca
      781 attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat agaataacat
      841 ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc tcggttctat
      901 cgaaaacgcg cctgctagcg ccaccatgaa gaccatcatt gccctgagct acatcttctg
      961 cctggtgttc gctgcggccg ccgtgatcca cgtgaccaaa gaagtgaaag aggtcgccac
     1021 actgagctgc ggccacaacg tttcagtgga agaactggcc cagaccagga tctactggca
     1081 gaaagaaaag aaaatggtgc tgaccatgat gagcggcgac atgaacatct ggcccgagta
     1141 caagaaccgg accatcttcg acatcaccaa caacctgagc atcgtgattc tggccctgag
     1201 gccttctgat gagggcacct atgagtgcgt ggtgctgaag tacgagaagg acgccttcaa
     1261 gcgggaacac ctggccgaag tgacactgag cgtgaaggcc gactttccca cacctagcat
     1321 cagcgacttc gagatcccca ccagcaacat cagacggatc atctgtagca ccagcggcgg
     1381 ctttccagag cctcatctgt cttggctgga aaacggcgag gaactgaacg ccatcaacac
     1441 caccgtgtct caggaccccg agacagagct gtatgccgtg tccagcaagc tggacttcaa
     1501 catgaccacc aaccacagct tcatgtgcct gattaagtac ggccacctga gagtgaacca
     1561 gaccttcaac tggaatacca ccaagcaaga gcacttcccc gacaatggcg cgccgggcgg
     1621 cgacaagacc cacacctgcc ccccctgccc agctccagaa ctgctgggag ggccttccgt
     1681 gttcctgttc ccccccaagc ccaaggacac cctgatgatc agcaggaccc ccgaggtgac
     1741 ctgcgtggtg gtggccgtgt cccacgagga cccagaggtg aagttcaact ggtacgtgga
     1801 cggcgtggag gtgcacaacg ccaagaccaa gcccagagag gagcagtaca acagcaccta
     1861 cagggtggtg tccgtgctga ccgtgctgca ccaggactgg ctgaacggca aagaatacaa
     1921 gtgcaaagtc tccaacaagg ccctggctgc cccaatcgaa aagacaatca gcaaggccaa
     1981 gggccagcca cgggagcccc aggtgtacac cctgcccccc agccgggagg agatgaccaa
     2041 gaaccaggtg tccctgacct gtctggtgaa gggcttctac cccagcgata tcgccgtgga
     2101 gtgggagagc aacggccagc ccgagaacaa ctacaagacc acccccccag tgctggacag
     2161 cgacggcagc ttcttcctgt acagcaagct gaccgtggac aagtccaggt ggcagcaggg
     2221 caacgtgttc agctgcagcg tgatgcacga ggccctgcac aaccactaca cccagaagtc
     2281 cctgagcctg agccccggcg gcagcggtct taatgatatt ttcgaggctc agaagattga
     2341 gtggcatgaa caccaccacc atcaccacta aggccggcct gtagtcactg gcacaacgcg
     2401 ttacccggta agccaatcgg gtatacacgg tcgtcatact gcagacaggg ttcttctact
     2461 ttgcaagata gtcttgagta gtaaaataaa tagatagaga aaatcagcct cgactgtgcc
     2521 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     2581 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcacaac actcaaccct
     2641 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     2701 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
     2761 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2821 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     2881 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     2941 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     3001 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     3061 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc
     3121 acgtgttgac aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag
     3181 gaactaaacc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc
     3241 cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt
     3301 cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt
     3361 cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac
     3421 cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga
     3481 gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg
     3541 gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa
     3601 gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc
     3661 cgccttcctg gagacatccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac
     3721 cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc
     3781 cggtgcctga cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     3841 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     3901 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     3961 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     4021 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     4081 gtaatcatgg tcattaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     4141 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     4201 taccatctgg ccccagcgct gcgatgatac cgcgagaacc acgctcaccg gctccggatt
     4261 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     4321 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     4381 atagtttgcg caacgttgtt gccatcgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     4441 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     4501 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     4561 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     4621 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     4681 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     4741 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     4801 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     4861 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     4921 gaataagggc gacacggaaa tgttgaatac tcatattctt cctttttcaa tattattgaa
     4981 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     5041 aacaaatagg ggtcagtgtt acaaccaatt aaccaattct gaacattatc gcgagcccat
     5101 ttatacctga atatggctca taacacccct tgctcatgac caaaatccct taacgtgagt
     5161 tacgcgcgcg tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     5221 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     5281 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     5341 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttagccc accacttcaa
     5401 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     5461 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     5521 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     5581 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     5641 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     5701 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     5761 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     5821 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     5881 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     5941 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaaggcg agagtaggga
     6001 actgccaggc atcaaactaa gcagaaggcc cctgacggat ggcctttttg cgtttctaca
     6061 aactctttct gtgttgtaaa acgacggcca gtcttaagct cgggccccct gggcggttct
     6121 gataacgagt aatcgttaat ccgcaaataa cgtaaaaacc cgcttcggcg ggttttttta
     6181 tggggggagt ttagggaaag agcatttgtc agaatattta agggcgcctg tcactttgct
     6241 tgatatatga gaattattta accttataaa tgagaaaaaa gcaacgcact ttaaataaga
     6301 tacgttgctt tttcgattga tgaacaccta taattaaact attcatctat tatttatgat
     6361 tttttgtata tacaatattt ctagtttgtt aaagagaatt aagaaaataa atctcgaaaa
     6421 taataaaggg aaaatcagtt tttgatatca aaattataca tgtcaacgat aatacaaaat
     6481 ataatacaaa ctataagatg ttatcagtat ttattatcat ttagaataaa ttttgtgtcg
     6541 cccttaattg tgagcggata acaattacga gcttcatgca cagtggcgtt gactcaatat
     6601 tggccattag ccatattatt cattggttat atagcataaa tcaatattgg ctattggcca
     6661 ttgcatacgt tgtatctata tcataatatg tacatttata ttggctcatg tccaatatga
     6721 ccgccatgtt
//
