LOCUS       sequence-421007-        6967 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 6967)
  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 6967)
  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-MICB-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6967
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(53..73)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        58..387
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      238..373
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     300..319
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        435..482
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             501..1100
                     /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(501..520)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     997..1017
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1230..1351
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1267..1286)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1321..1340
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(1404..2195,2196..2264))
                     /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     2027..2046
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2265..2356)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2398..2417
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2423..2442
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2528..3116
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3017..3036
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3270..3287
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     3385..3402
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3386..3402
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3858..3874
                     /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        4041..4420
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4421..4624
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4575..4595
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4621..4645
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        4855..4873
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4855..4872
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     4886..4905
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             6583..6627
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6628..6645
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6785..6802)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6791..6902
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
       61 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
      121 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
      181 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
      241 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
      301 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
      361 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
      421 tctgatcagc acgtgttgac aattaatcat cggcatagta tatcggcata gtataatacg
      481 acaaggtgag gaactaaacc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg
      541 acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc
      601 gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc
      661 tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg
      721 cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc
      781 gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg
      841 cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc
      901 accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg
      961 ccggggtgcc cgccttcctg gagacatccg cgccccgcaa cctccccttc tacgagcggc
     1021 tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga
     1081 cccgcaagcc cggtgcctga cacgtgctac gagatttcga ttccaccgcc gccttctatg
     1141 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     1201 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca
     1261 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     1321 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct
     1381 agagcttggc gtaatcatgg tcattaccaa tgcttaatca gtgaggcacc tatctcagcg
     1441 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     1501 cgggagggct taccatctgg ccccagcgct gcgatgatac cgcgagaacc acgctcaccg
     1561 gctccggatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     1621 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     1681 tcgccagtta atagtttgcg caacgttgtt gccatcgcta caggcatcgt ggtgtcacgc
     1741 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     1801 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     1861 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     1921 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     1981 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     2041 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     2101 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     2161 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     2221 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatattctt cctttttcaa
     2281 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     2341 tagaaaaata aacaaatagg ggtcagtgtt acaaccaatt aaccaattct gaacattatc
     2401 gcgagcccat ttatacctga atatggctca taacacccct tgctcatgac caaaatccct
     2461 taacgtgagt tacgcgcgcg tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     2521 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     2581 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     2641 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttagccc
     2701 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     2761 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     2821 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     2881 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     2941 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     3001 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     3061 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     3121 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     3181 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     3241 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaaggcg
     3301 agagtaggga actgccaggc atcaaactaa gcagaaggcc cctgacggat ggcctttttg
     3361 cgtttctaca aactctttct gtgttgtaaa acgacggcca gtcttaagct cgggccccct
     3421 gggcggttct gataacgagt aatcgttaat ccgcaaataa cgtaaaaacc cgcttcggcg
     3481 ggttttttta tggggggagt ttagggaaag agcatttgtc agaatattta agggcgcctg
     3541 tcactttgct tgatatatga gaattattta accttataaa tgagaaaaaa gcaacgcact
     3601 ttaaataaga tacgttgctt tttcgattga tgaacaccta taattaaact attcatctat
     3661 tatttatgat tttttgtata tacaatattt ctagtttgtt aaagagaatt aagaaaataa
     3721 atctcgaaaa taataaaggg aaaatcagtt tttgatatca aaattataca tgtcaacgat
     3781 aatacaaaat ataatacaaa ctataagatg ttatcagtat ttattatcat ttagaataaa
     3841 ttttgtgtcg cccttaattg tgagcggata acaattacga gcttcatgca cagtggcgtt
     3901 gactcaatat tggccattag ccatattatt cattggttat atagcataaa tcaatattgg
     3961 ctattggcca ttgcatacgt tgtatctata tcataatatg tacatttata ttggctcatg
     4021 tccaatatga ccgccatgtt ggcattgatt attgactagt tattaatagt aatcaattac
     4081 ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg
     4141 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
     4201 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
     4261 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa
     4321 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac
     4381 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta
     4441 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga
     4501 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa
     4561 ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag
     4621 agctcgttta gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca
     4681 tagaagacac cgggaccgat ccagcctccg cggccgggaa cggtgcattg gaacgcggat
     4741 tccccgtgcc aagagtgacg taagtaccgc ctatagagtc tataggccca cccccttggc
     4801 ttcgttagaa cgcggctaca attaatacat aaccttatgt atcatacaca tacgatttag
     4861 gtgacactat agaataacat ccactttgcc tttctctcca caggtgtcca ctcccaggtc
     4921 caactgcacc tcggttctat cgaaaacgcg cctgctagcg ccaccatgaa gaccatcatt
     4981 gccctgagct acatcttctg cctggtgttc gctgcggccg ccgctgaacc tcactctctg
     5041 agatacaacc tgatggtgct gagccaggac gagagcgtgc agtctggatt tctggccgaa
     5101 ggacacctgg acggccagcc tttcctgaga tacgacagac agaagcggag agccaagcct
     5161 caaggacagt gggctgaaga tgtgctgggc gccaagacct gggataccga gacagaggat
     5221 ctgaccgaga acggccagga cctgagaaga accctgacac acatcaagga ccagaaaggc
     5281 ggcctgcaca gcctgcaaga gatcagagtg tgcgagatcc acgaggacag cagcacaaga
     5341 ggcagccggc acttttacta cgacggcgag ctgttcctga gccagaacct ggaaacccaa
     5401 gagtccaccg tgcctcagtc tagcagagcc cagacactgg ccatgaacgt gaccaatttc
     5461 tggaaagagg acgccatgaa gaccaagaca cactacagag ccatgcaggc cgactgcctg
     5521 cagaagctgc agagatatct gaagtccggc gtggccatca gacggacagt gcctccaatg
     5581 gtcaacgtga cctgttctga ggtgtccgag ggcaacatca ccgtgacatg cagagccagc
     5641 agcttctacc ccagaaatat caccctgacc tggcggcagg atggcgtgtc cctgtctcat
     5701 aatacccagc agtggggcga cgtgctgcct gatggcaatg gcacatatca gacctgggtc
     5761 gccaccagaa tccggcaggg cgaagaacag cggttcacct gttacatgga acacagcggc
     5821 aaccacggca cacaccctgt gccatctgga aaagtgctgg tgctgcagtc ccagcggaca
     5881 gatggcgcgc cgggcggcga caagacccac acctgccccc cctgcccagc tccagaactg
     5941 ctgggagggc cttccgtgtt cctgttcccc cccaagccca aggacaccct gatgatcagc
     6001 aggacccccg aggtgacctg cgtggtggtg gccgtgtccc acgaggaccc agaggtgaag
     6061 ttcaactggt acgtggacgg cgtggaggtg cacaacgcca agaccaagcc cagagaggag
     6121 cagtacaaca gcacctacag ggtggtgtcc gtgctgaccg tgctgcacca ggactggctg
     6181 aacggcaaag aatacaagtg caaagtctcc aacaaggccc tggctgcccc aatcgaaaag
     6241 acaatcagca aggccaaggg ccagccacgg gagccccagg tgtacaccct gccccccagc
     6301 cgggaggaga tgaccaagaa ccaggtgtcc ctgacctgtc tggtgaaggg cttctacccc
     6361 agcgatatcg ccgtggagtg ggagagcaac ggccagcccg agaacaacta caagaccacc
     6421 cccccagtgc tggacagcga cggcagcttc ttcctgtaca gcaagctgac cgtggacaag
     6481 tccaggtggc agcagggcaa cgtgttcagc tgcagcgtga tgcacgaggc cctgcacaac
     6541 cactacaccc agaagtccct gagcctgagc cccggcggca gcggtcttaa tgatattttc
     6601 gaggctcaga agattgagtg gcatgaacac caccaccatc accactaagg ccggcctgta
     6661 gtcactggca caacgcgtta cccggtaagc caatcgggta tacacggtcg tcatactgca
     6721 gacagggttc ttctactttg caagatagtc ttgagtagta aaataaatag atagagaaaa
     6781 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     6841 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     6901 tcacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     6961 cggccta
//
