LOCUS       sequence-461172-        7921 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to COMP5AP-AviTag-9xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7921)
  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 7921)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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-JAML-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7921
                     /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             3571..3594
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5254..5298
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5299..5325
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(5465..5482)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5471..5582
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5700..5720)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5705..6034
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5885..6020
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5947..5966
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6082..6129
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6148..6747
                     /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(6148..6167)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6644..6664
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6877..6998
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6914..6933)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6968..6987
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7051..7842,7843..7911))
                     /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     7674..7693
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7912..7921,1..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 gaacgatctg
     2761 aatgtgtctc cacctgagct gaccgtgcac gtgggagatt ctgctctgat gggctgtgtg
     2821 ttccagagca ccgaggacaa gtgcatcttc aagatcgact ggacactgag ccctggcgag
     2881 cacgccaagg atgagtacgt gctgtactac tactccaacc tgagcgtgcc catcggccgg
     2941 ttccagaata gagtgcacct gatgggcgac atcctgtgca atgatggaag cctgctgctg
     3001 caggacgtgc aagaagccga tcagggcacc tacatctgcg agatcagact gaagggcgag
     3061 agccaggtgt tcaagaaagc cgtggtgctg cacgtgctgc ccgaggaacc taaagaactg
     3121 atggtgcacg ttggcggcct gatccagatg ggatgtgtct ttcagtccac cgaagtgaag
     3181 cacgtgacca aggtggaatg gatctttagc ggcagacggg ccaaagagga aatcgtcttt
     3241 cggtactacc acaagctgcg gatgagcgtg gaatactctc agagctgggg ccactttcag
     3301 aacagagtga atctcgtggg cgatatcttc cggaacgacg gctctatcat gctgcagggc
     3361 gtcagagaga gcgacggcgg caattacaca tgtagcatcc acctgggcaa cctcgtgttc
     3421 aaaaagacaa tcgtgctgca tgtgtcccca gaagaaccca gaacactggt cacaccagct
     3481 gctctgaggc ctctggtgct cggcggaaat caacttggcg cgccggaaga tcaagttgat
     3541 ccaagattga ttgatggcaa gggcggatct ctggaggtgc tgttccaggg accaggcgga
     3601 agcggcaatc ccggcggcga tctggccccc cagatgctgc gcgagctgca ggagaccaat
     3661 gccgccctgc aggacgtgcg cgagctgctg cgccaccgcg tgaaggagat caccttcctg
     3721 aagaacaccg tgatggagtg cgacgcctgc ggtatgcagc cctctagagg ctccggctcc
     3781 atcatcccag ttgaggagga gaacccggac ttctggaacc gcgaggcagc cgaggccctg
     3841 ggtgccgcca agaagctgca gcctgcacag acagccgcca agaacctcat catcttcctg
     3901 ggcgatggga tgggggtgtc tacggtgaca gctgccagga tcctaaaagg gcagaagaag
     3961 gacaaactgg ggcctgagat acccctggcc atggaccgct tcccatatgt ggctctgtcc
     4021 aagacataca atgtagacaa acatgtgcca gacagtggag ccacagccac ggcctacctg
     4081 tgcggggtca agggcaactt ccagaccatt ggcttgagtg cagccgcccg ctttaaccag
     4141 tgcaacacga cacgcggcaa cgaggtcatc tccgtgatga atcgggccaa gaaagcaggg
     4201 aagtcagtgg gagtggtaac caccacacga gtgcagcacg cctcgccagc cggcacctac
     4261 gcccacacgg tgaaccgcaa ctggtactcg gacgccgacg tgcctgcctc ggcccgccag
     4321 gaggggtgcc aggacatcgc tacgcagctc atctccaaca tggacattga cgtgatccta
     4381 ggtggaggcc gaaagtacat gtttcgcatg ggaaccccag accctgagta cccagatgac
     4441 tacagccaag gtgggaccag gctggacggg aagaatctgg tgcaggaatg gctggcgaag
     4501 cgccagggtg cccggtatgt gtggaaccgc actgagctca tgcaggcttc cctggacccg
     4561 tctgtgaccc atctcatggg tctctttgag cctggagaca tgaaatacga gatccaccga
     4621 gactccacac tggacccctc cctgatggag atgacagagg ctgccctgcg cctgctgagc
     4681 aggaaccccc gcggcttctt cctcttcgtg gagggtggtc gcatcgacca tggtcatcat
     4741 gaaagcaggg cttaccgggc actgactgag acgatcatgt tcgacgacgc cattgagagg
     4801 gcgggccagc tcaccagcga ggaggacacg ctgagcctcg tcactgccga ccactcccac
     4861 gtcttctcct tcggaggcta ccccctgcga gggagctcca tcttcgggct ggcccctggc
     4921 aaggcccggg acaggaaggc ctacacggtc ctcctatacg gaaacggtcc aggctatgtg
     4981 ctcaaggacg gcgcccggcc ggatgttacc gagagcgaga gcgggagccc cgagtatcgg
     5041 cagcagtcag cagtgcccct ggacgaagag acccacgcag gcgaggacgt ggcggtgttc
     5101 gcgcgcggcc cgcaggcgca cctggttcac ggcgtgcagg agcagacctt catagcgcac
     5161 gtcatggcct tcgccgcctg cctggagccc tacaccgcct gcgacctggc gccccccgcc
     5221 ggcaccaccg acgccgcgca cccgggtggc ggaggactga acgacatctt cgaggctcag
     5281 aaaatcgaat ggcacgaaca tcaccatcac catcaccatc accattaagg ccggcctgta
     5341 gtcactggca caacgcgtta cccggtaagc caatcgggta tacacggtcg tcatactgca
     5401 gacagggttc ttctactttg caagatagtc ttgagtagta aaataaatag atagagaaaa
     5461 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     5521 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     5581 tcacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     5641 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg
     5701 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     5761 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     5821 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     5881 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     5941 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     6001 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     6061 tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat cggcatagta
     6121 taatacgaca aggtgaggaa ctaaaccatg accgagtaca agcccacggt gcgcctcgcc
     6181 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc
     6241 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa
     6301 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc
     6361 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc
     6421 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc
     6481 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg
     6541 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc
     6601 gagcgcgccg gggtgcccgc cttcctggag acatccgcgc cccgcaacct ccccttctac
     6661 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg
     6721 tgcatgaccc gcaagcccgg tgcctgacac gtgctacgag atttcgattc caccgccgcc
     6781 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     6841 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
     6901 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     6961 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
     7021 tctagctaga gcttggcgta atcatggtca ttaccaatgc ttaatcagtg aggcacctat
     7081 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     7141 tacgatacgg gagggcttac catctggccc cagcgctgcg atgataccgc gagaaccacg
     7201 ctcaccggct ccggatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     7261 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     7321 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc atcgctacag gcatcgtggt
     7381 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     7441 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     7501 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     7561 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     7621 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     7681 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     7741 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     7801 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     7861 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tattcttcct
     7921 t
//
