LOCUS       sequence-74805-e        6003 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of S. pyogenes SrtA at the plasma membrane in mammalian 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6003)
  AUTHORS   Strijbis K, Spooner E, Ploegh HL
  TITLE     Protein ligation in living cells using sortase.
  JOURNAL   Traffic. 2012 Jun;13(6):780-9. doi: 
            10.1111/j.1600-0854.2012.01345.x. Epub 2012 Mar 23.
  PUBMED    22348280
REFERENCE   2  (bases 1 to 6003)
  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: 
            "10.1111/j.1600-0854.2012.01345.x"; journalName: "Traffic"; date: 
            "2012-06"; volume: "13"; issue: "6"; pages: "780-9"
COMMENT     Sequence Label: pcDNA3.1(-)-PM-Cpalm-HA-SrtA (pKS37)
FEATURES             Location/Qualifiers
     source          1..6003
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      921..930
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             978..1004
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(1597..1614)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1603..1827
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1873..2301
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1960..1979)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2170..2191
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2310..2330)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2315..2644
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2495..2630
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2557..2576
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2711..3505
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2765..2784)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3375..3394
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3679..3800
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3716..3735)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3770..3789
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3849..3865)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3849..3865)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3862..3884)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3873..3889
                     /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)."
     promoter        complement(join(3897..3903,3904..3921,3922..3927))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3942..3963
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4080..4097)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4251..4836)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4331..4350)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5007..5798,5799..5867))
                     /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     5630..5649
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5868..5972)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga gccaccatgg gatgtataaa atcaaaaggg aaagacagcg
      961 cgggagcaat cgatgcctac ccatacgacg taccagacta cgcacatagt gctggtggat
     1021 ccgtcttgca agcacaaatg gcggctcagc aacttcctgt tatagggggc attgccatac
     1081 cagagcttgg cattaattta ccaattttta aaggtttagg aaatactgag cttatttatg
     1141 gcgcaggaac gatgaaagaa gaacaagtta tgggaggaga aaataattat tctcttgcca
     1201 gtcatcatat ttttggaatt acaggttcat ctcaaatgct cttttcgccg cttgaaagag
     1261 cacaaaatgg gatgtccatc tatttaacag ataaagaaaa aatttacgaa tacatcataa
     1321 aagatgtttt cacggtagct cctgaacgcg ttgatgttat cgatgataca gctggtctca
     1381 aagaagtgac tttagtgact tgtacagata tcgaagcaac agaacgtatt attgtcaaag
     1441 gagaactaaa aacagaatac gactttgata aagcgcccgc cgatgtattg aaagctttta
     1501 atcattctta taaccaagta tctacctagg aattccacca cactggacta gtggatccga
     1561 gctcggtacc aagcttaagt ttaaaccgct gatcagcctc gactgtgcct tctagttgcc
     1621 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     1681 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     1741 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     1801 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     1861 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     1921 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     1981 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     2041 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     2101 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     2161 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     2221 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     2281 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     2341 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     2401 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     2461 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     2521 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     2581 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     2641 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcaagaga caggatgagg
     2701 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     2761 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     2821 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     2881 gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     2941 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     3001 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     3061 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     3121 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     3181 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg
     3241 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     3301 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     3361 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     3421 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     3481 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     3541 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     3601 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     3661 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     3721 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     3781 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     3841 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     3901 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     3961 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     4021 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4081 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4141 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4201 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4261 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4321 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4381 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4441 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4501 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4561 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4621 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     4681 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     4741 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggttt ttttgtttgc
     4801 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     4861 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     4921 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     4981 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     5041 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     5101 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     5161 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     5221 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     5281 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     5341 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     5401 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     5461 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     5521 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     5581 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     5641 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     5701 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     5761 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     5821 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     5881 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     5941 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     6001 gtc
//
