LOCUS       sequence-303071-        8524 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein partial 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 8524)
  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 8524)
  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-NOTCH2(Trunc2)-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8524
                     /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             5866..5910
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5911..5928
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6068..6085)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6074..6185
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6303..6323)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6308..6637
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6488..6623
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6550..6569
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6685..6732
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6751..7350
                     /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(6751..6770)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7247..7267
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7480..7601
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7517..7536)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7571..7590
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7654..8445,8446..8514))
                     /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     8277..8296
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8515..8524,1..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 gcaatgtaga
     2761 gatggctacg agccctgcgt gaacgaaggg atgtgtgtga cctaccacaa cggcaccggc
     2821 tactgcaagt gccctgaggg atttctgggc gagtactgcc agcacagaga tccctgcgag
     2881 aagaacagat gccagaacgg cggaacctgt gtggcccagg ctatgctggg aaaagccacc
     2941 tgtagatgcg ccagcggctt taccggcgag gattgtcagt acagcaccag ccatccttgc
     3001 ttcgtgtccc ggccttgtct gaatggcggc acctgtcaca tgctgagcag agacacctac
     3061 gagtgtacct gccaagtggg cttcaccggc aaagaatgcc agtggaccga tgcctgcctg
     3121 tctcaccctt gtgccaatgg cagcacatgt accaccgtgg ccaaccagtt cagctgcaag
     3181 tgtctgaccg gcttcacagg ccagaaatgc gagacagacg tgaacgagtg cgacatccct
     3241 ggccactgtc agcatggcgg aacatgcctg aatctgcccg gcagctacca gtgccagtgt
     3301 cctcaaggct tcactggaca gtactgcgac agcctgtacg tgccctgtgc tccttctcct
     3361 tgtgtgaacg gcgggacatg tagacagacc ggcgacttca ccttcgagtg caactgcctg
     3421 ccaggcttcg agggcagcac ttgcgagaga aacatcgacg actgccccaa ccaccggtgc
     3481 caaaatggcg gagtgtgtgt ggatggcgtg aacacctaca actgcagatg ccctcctcag
     3541 tggacaggcc agttctgcac cgaggatgtg gatgagtgtc tgctgcagcc taacgcctgt
     3601 caaaacggcg gcacttgcgc caacagaaat ggcggctatg gctgcgtgtg tgtcaacgga
     3661 tggtccggcg acgattgcag cgagaacatc gatgattgcg ccttcgccag ctgcacccct
     3721 ggctctacct gtatcgatag agtggccagc ttctcctgca tgtgtcccga aggcaaagcc
     3781 ggcctgctgt gccatctgga tgatgcctgc atcagcaacc cctgtcacaa aggcgccctg
     3841 tgcgacacca atcctctgaa cggccagtac atctgcacct gtcctcaggg ctataagggc
     3901 gccgactgta cagaggacgt ggacgaatgt gccatggcca acagcaaccc ttgtgaacac
     3961 gccgggaaat gtgtgaacac cgacggcgcc tttcactgcg agtgcctgaa aggatacgct
     4021 ggccccagat gcgagatgga catcaacgag tgccacagcg acccctgcca gaatgatgcc
     4081 acctgtctgg ataagatcgg cggcttcact tgcctgtgca tgcctggctt taagggcgtg
     4141 cactgcgagc tggaaatcaa tgaatgtcaa agcaacccgt gcgtgaacaa tggccagtgc
     4201 gtggacaaag tgaacaggtt ccagtgtctg tgcccacctg gattcaccgg acctgtgtgc
     4261 cagatcgaca ttgatgactg ctccagcaca ccctgcctga acggcgccaa gtgtatcgat
     4321 caccccaacg gctatgagtg tcagtgtgcc acaggcttta caggcgtgct gtgcgaagag
     4381 aatatcgaca actgcgaccc cgatccttgc caccacggac agtgtcagga tggcatcgac
     4441 agctacacct gtatctgcaa ccccggctac atgggcgcca tctgcagcga tcagatcgat
     4501 gagtgctaca gcagcccatg tctgaacgac ggcagatgca tcgacctggt caatggctat
     4561 cagtgcaatt gccagcctgg caccagcgga gtgaactgcg agatcaactt cgatgactgt
     4621 gccagcaatc cctgtatcca cggcatctgt atggacggca tcaaccggta cagctgtgtg
     4681 tgcagcccag ggtttaccgg ccagagatgc aacattgata tcgacgagtg cgcctccaat
     4741 ccatgcagaa agggcgccac atgcatcaac ggcgtgaacg gcttccggtg catctgtcct
     4801 gaaggacccc accatcctag ctgctacagc caagtgaacg aatgcctgag caacccatgc
     4861 atccacggaa actgtacagg cggcctgagc ggctacaagt gcctgtgtga tgctggctgg
     4921 gtcggaatca actgcgaggt ggacaagaat gagtgcctgt ctaatccctg tcagaatggt
     4981 ggcacatgtg acaatctcgt gaatggctac aggtgcacat gcaagaaggg cttcaagggg
     5041 tacaactgcc aagtcaacat tgacgagtgt gcttctaacc cctgcctcaa ccagggcacc
     5101 tgtttcgacg acatcagcgg ctatacctgc cactgcgtgc tgccttacac cggcaagaac
     5161 tgtcaaggcg cgccgggcgg cgacaagacc cacacctgcc ccccctgccc agctccagaa
     5221 ctgctgggag ggccttccgt gttcctgttc ccccccaagc ccaaggacac cctgatgatc
     5281 agcaggaccc ccgaggtgac ctgcgtggtg gtggccgtgt cccacgagga cccagaggtg
     5341 aagttcaact ggtacgtgga cggcgtggag gtgcacaacg ccaagaccaa gcccagagag
     5401 gagcagtaca acagcaccta cagggtggtg tccgtgctga ccgtgctgca ccaggactgg
     5461 ctgaacggca aagaatacaa gtgcaaagtc tccaacaagg ccctggctgc cccaatcgaa
     5521 aagacaatca gcaaggccaa gggccagcca cgggagcccc aggtgtacac cctgcccccc
     5581 agccgggagg agatgaccaa gaaccaggtg tccctgacct gtctggtgaa gggcttctac
     5641 cccagcgata tcgccgtgga gtgggagagc aacggccagc ccgagaacaa ctacaagacc
     5701 acccccccag tgctggacag cgacggcagc ttcttcctgt acagcaagct gaccgtggac
     5761 aagtccaggt ggcagcaggg caacgtgttc agctgcagcg tgatgcacga ggccctgcac
     5821 aaccactaca cccagaagtc cctgagcctg agccccggcg gcagcggtct taatgatatt
     5881 ttcgaggctc agaagattga gtggcatgaa caccaccacc atcaccacta aggccggcct
     5941 gtagtcactg gcacaacgcg ttacccggta agccaatcgg gtatacacgg tcgtcatact
     6001 gcagacaggg ttcttctact ttgcaagata gtcttgagta gtaaaataaa tagatagaga
     6061 aaatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     6121 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     6181 gcatcacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     6241 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct
     6301 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     6361 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     6421 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     6481 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     6541 aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta
     6601 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc
     6661 cattttcgga tctgatcagc acgtgttgac aattaatcat cggcatagta tatcggcata
     6721 gtataatacg acaaggtgag gaactaaacc atgaccgagt acaagcccac ggtgcgcctc
     6781 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     6841 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     6901 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     6961 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     7021 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa
     7081 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc
     7141 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg
     7201 gccgagcgcg ccggggtgcc cgccttcctg gagacatccg cgccccgcaa cctccccttc
     7261 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     7321 tggtgcatga cccgcaagcc cggtgcctga cacgtgctac gagatttcga ttccaccgcc
     7381 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     7441 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     7501 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     7561 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     7621 acctctagct agagcttggc gtaatcatgg tcattaccaa tgcttaatca gtgaggcacc
     7681 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     7741 aactacgata cgggagggct taccatctgg ccccagcgct gcgatgatac cgcgagaacc
     7801 acgctcaccg gctccggatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     7861 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     7921 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccatcgcta caggcatcgt
     7981 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     8041 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     8101 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     8161 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     8221 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     8281 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     8341 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     8401 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     8461 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatattctt
     8521 cctt
//
