LOCUS       sequence-368527-        6589 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 6589)
  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 6589)
  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-CD8A-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6589
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     213..233
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    351..462
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     451..468
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(608..625)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(626..670)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(1970..1989)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(2002..2020)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2003..2020)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2230..2254)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2251..2454)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2280..2300)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2455..2834
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    3001..3017
                     /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(3473..3490)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3473..3489)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3588..3605)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3759..4347)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3839..3858)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4433..4452)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4458..4477)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4519..4610
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4611..4679,4680..5471)
                     /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(4829..4848)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    5524..5645
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5535..5554)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5589..5608
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5775..6374)
                     /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(5858..5878)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6355..6374
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6393..6440)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(join(6488..6589,1..228))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6502..6589,1..48)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6556..6575)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 gccatggggc ggagaatggg cggaactggg cggagttagg ggcgggatgg gcggagttag
       61 gggcgggact atggttgctg actaattgag atgcatgctt tgcatacttc tgcctgctgg
      121 ggagcctggg gactttccac acctggttgc tgactaattg agatgcatgc tttgcatact
      181 tctgcctgct ggggagcctg gggactttcc acaccctaac tgacacacat tccacagaat
      241 taattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc
      301 aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt gatgcaattt
      361 cctcatttta ttaggaaagg acagtgggag tggcaccttc cagggtcaag gaaggcacgg
      421 gggaggggca aacaacagat ggctggcaac tagaaggcac agtcgaggct gattttctct
      481 atctatttat tttactactc aagactatct tgcaaagtag aagaaccctg tctgcagtat
      541 gacgaccgtg tatacccgat tggcttaccg ggtaacgcgt tgtgccagtg actacaggcc
      601 ggccttagtg gtgatggtgg tggtgttcat gccactcaat cttctgagcc tcgaaaatat
      661 cattaagacc gctgccgccg gggctcaggc tcagggactt ctgggtgtag tggttgtgca
      721 gggcctcgtg catcacgctg cagctgaaca cgttgccctg ctgccacctg gacttgtcca
      781 cggtcagctt gctgtacagg aagaagctgc cgtcgctgtc cagcactggg ggggtggtct
      841 tgtagttgtt ctcgggctgg ccgttgctct cccactccac ggcgatatcg ctggggtaga
      901 agcccttcac cagacaggtc agggacacct ggttcttggt catctcctcc cggctggggg
      961 gcagggtgta cacctggggc tcccgtggct ggcccttggc cttgctgatt gtcttttcga
     1021 ttggggcagc cagggccttg ttggagactt tgcacttgta ttctttgccg ttcagccagt
     1081 cctggtgcag cacggtcagc acggacacca ccctgtaggt gctgttgtac tgctcctctc
     1141 tgggcttggt cttggcgttg tgcacctcca cgccgtccac gtaccagttg aacttcacct
     1201 ctgggtcctc gtgggacacg gccaccacca cgcaggtcac ctcgggggtc ctgctgatca
     1261 tcagggtgtc cttgggcttg ggggggaaca ggaacacgga aggccctccc agcagttctg
     1321 gagctgggca gggggggcag gtgtgggtct tgtcgccgcc cggcgcgcca tcacaggcga
     1381 agtccagtcc tcttgtatgc acggctccgc cagcagcagg tctacaagct tctggcctca
     1441 gagacagagg ctggctggca attgtaggag ctggtgtagg tggtctagga gcgggtgttg
     1501 ttgtaggctt ggcgggcaga aacacgggca cgaagtggct gaagtacatg atgctgttgc
     1561 tcagggcgct gcagaagtag tagccctcgt tctctcttct gaagtcgctc agggtcagca
     1621 cgaaggtgtc gcccagtctc ttgccagaga atctctgtgt atccaggcct tcggcggcct
     1681 taggcttgtt ctggctcagg tacagcagaa aggtgggaga agcggcagca cctctaggct
     1741 gaaacagcca gctgcagccg cttgtaggat tgctcagcag cacctggcac ttcagttcca
     1801 ctgtctctcc caggttccag gttctgtcca gaggggacac tctaaactga gaggcggccg
     1861 cagcgaacac caggcagaag atgtagctca gggcaatgat ggtcttcatg gtggcgctag
     1921 caggcgcgtt ttcgatagaa ccgaggtgca gttggacctg ggagtggaca cctgtggaga
     1981 gaaaggcaaa gtggatgtta ttctatagtg tcacctaaat cgtatgtgta tgatacataa
     2041 ggttatgtat taattgtagc cgcgttctaa cgaagccaag ggggtgggcc tatagactct
     2101 ataggcggta cttacgtcac tcttggcacg gggaatccgc gttccaatgc accgttcccg
     2161 gccgcggagg ctggatcggt cccggtgtct tctatggagg tcaaaacagc gtggatggcg
     2221 tctccaggcg atctgacggt tcactaaacg agctctgctt atatagacct cccaccgtac
     2281 acgcctaccg cccatttgcg tcaatggggc ggagttgtta cgacattttg gaaagtcccg
     2341 ttgattttgg tgccaaaaca aactcccatt gacgtcaatg gggtggagac ttggaaatcc
     2401 ccgtgagtca aaccgctatc cacgcccatt gatgtactgc caaaaccgca tcaccatggt
     2461 aatagcgatg actaatacgt agatgtactg ccaagtagga aagtcccata aggtcatgta
     2521 ctgggcataa tgccaggcgg gccatttacc gtcattgacg tcaatagggg gcgtacttgg
     2581 catatgatac acttgatgta ctgccaagtg ggcagtttac cgtaaatact ccacccattg
     2641 acgtcaatgg aaagtcccta ttggcgttac tatgggaaca tacgtcatta ttgacgtcaa
     2701 tgggcggggg tcgttgggcg gtcagccagg cgggccattt accgtaagtt atgtaacgcg
     2761 gaactccata tatgggctat gaactaatga ccccgtaatt gattactatt aataactagt
     2821 caataatcaa tgccaacatg gcggtcatat tggacatgag ccaatataaa tgtacatatt
     2881 atgatataga tacaacgtat gcaatggcca atagccaata ttgatttatg ctatataacc
     2941 aatgaataat atggctaatg gccaatattg agtcaacgcc actgtgcatg aagctcgtaa
     3001 ttgttatccg ctcacaatta agggcgacac aaaatttatt ctaaatgata ataaatactg
     3061 ataacatctt atagtttgta ttatattttg tattatcgtt gacatgtata attttgatat
     3121 caaaaactga ttttcccttt attattttcg agatttattt tcttaattct ctttaacaaa
     3181 ctagaaatat tgtatataca aaaaatcata aataatagat gaatagttta attataggtg
     3241 ttcatcaatc gaaaaagcaa cgtatcttat ttaaagtgcg ttgctttttt ctcatttata
     3301 aggttaaata attctcatat atcaagcaaa gtgacaggcg cccttaaata ttctgacaaa
     3361 tgctctttcc ctaaactccc cccataaaaa aacccgccga agcgggtttt tacgttattt
     3421 gcggattaac gattactcgt tatcagaacc gcccaggggg cccgagctta agactggccg
     3481 tcgttttaca acacagaaag agtttgtaga aacgcaaaaa ggccatccgt caggggcctt
     3541 ctgcttagtt tgatgcctgg cagttcccta ctctcgcctt ccgcttcctc gctcactgac
     3601 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     3661 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     3721 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     3781 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     3841 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     3901 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     3961 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4021 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     4081 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     4141 tatgtaggcg gtgctacaga gttcttgaag tggtgggcta actacggcta cactagaaga
     4201 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     4261 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     4321 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     4381 gctcagtgga acgacgcgcg cgtaactcac gttaagggat tttggtcatg agcaaggggt
     4441 gttatgagcc atattcaggt ataaatgggc tcgcgataat gttcagaatt ggttaattgg
     4501 ttgtaacact gacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     4561 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagaat atgagtattc
     4621 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     4681 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     4741 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     4801 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     4861 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     4921 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     4981 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     5041 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     5101 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcga
     5161 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     5221 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     5281 cggctggctg gtttattgct gataaatccg gagccggtga gcgtggttct cgcggtatca
     5341 tcgcagcgct ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     5401 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     5461 agcattggta atgaccatga ttacgccaag ctctagctag aggtcgacgg tatacagaca
     5521 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     5581 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     5641 aagttggggt gggcgaagaa ctccagcatg agatccccgc gctggaggat catccagccg
     5701 gcgtcccgga aaacgattcc gaagcccaac ctttcataga aggcggcggt ggaatcgaaa
     5761 tctcgtagca cgtgtcaggc accgggcttg cgggtcatgc accaggtgcg cggtccttcg
     5821 ggcacctcga cgtcggcggt gacggtgaag ccgagccgct cgtagaaggg gaggttgcgg
     5881 ggcgcggatg tctccaggaa ggcgggcacc ccggcgcgct cggccgcctc cactccgggg
     5941 agcacgacgg cgctgcccag acccttgccc tggtggtcgg gcgagacgcc gacggtggcc
     6001 aggaaccacg cgggctcctt gggccggtgc ggcgccagga ggccttccat ctgttgctgc
     6061 gcggccagcc gggaaccgct caactcggcc atgcgcgggc cgatctcggc gaacaccgcc
     6121 cccgcttcga cgctctccgg cgtggtccag accgccaccg cggcgccgtc gtccgcgacc
     6181 cacaccttgc cgatgtcgag cccgacgcgc gtgaggaaga gttcttgcag ctcggtgacc
     6241 cgctcgatgt ggcggtccgg atcgacggtg tggcgcgtgg cggggtagtc ggcgaacgcg
     6301 gcggcgaggg tgcgtacggc cctggggacg tcgtcgcggg tggcgaggcg caccgtgggc
     6361 ttgtactcgg tcatggttta gttcctcacc ttgtcgtatt atactatgcc gatatactat
     6421 gccgatgatt aattgtcaac acgtgctgat cagatccgaa aatggatata caagctcccg
     6481 ggagcttttt gcaaaagcct aggcctccaa aaaagcctcc tcactacttc tggaatagct
     6541 cagaggcaga ggcggcctcg gcctctgcat aaataaaaaa aattagtca
//
