LOCUS       sequence-74804-e        6006 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of cytosolic S. pyogenes SrtA in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6006)
  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 6006)
  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(-)-HA-SrtAstrep (pKS77)
FEATURES             Location/Qualifiers
     source          1..6006
                     /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"
     CDS             939..965
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             975..992
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1600..1617)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1606..1830
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1876..2304
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1963..1982)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2173..2194
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2313..2333)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2318..2647
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2498..2633
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2560..2579
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2714..3508
                     /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(2768..2787)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3378..3397
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3682..3803
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3719..3738)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3773..3792
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3852..3868)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3852..3868)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3865..3887)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3876..3892
                     /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(3900..3906,3907..3924,3925..3930))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3945..3966
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4083..4100)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4254..4839)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4334..4353)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5010..5801,5802..5870))
                     /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     5633..5652
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5871..5975)
                     /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 gccaccatgc gcagcagcta cccatacgac gtaccagact
      961 acgcaagcag cggcctggtg ccgcgcggat ccgtcttgca agcacaaatg gcggctcagc
     1021 aacttcctgt tatagggggc attgccatac cagagcttgg cattaattta ccaattttta
     1081 aaggtttagg aaatactgag cttatttatg gcgcaggaac gatgaaagaa gaacaagtta
     1141 tgggaggaga aaataattat tctcttgcca gtcatcatat ttttggaatt acaggttcat
     1201 ctcaaatgct cttttcgccg cttgaaagag cacaaaatgg gatgtccatc tatttaacag
     1261 ataaagaaaa aatttacgaa tacatcataa aagatgtttt cacggtagct cctgaacgcg
     1321 ttgatgttat cgatgataca gctggtctca aagaagtgac tttagtgact tgtacagata
     1381 tcgaagcaac agaacgtatt attgtcaaag gagaactaaa aacagaatac gactttgata
     1441 aagcgcccgc cgatgtattg aaagctttta atcattctta taaccaagta tctacctagc
     1501 tcgagcggcc gccactgtgc tggatatctg cagaattcca ccacactgga ctagtggatc
     1561 cgagctcggt accaagctta agtttaaacc gctgatcagc ctcgactgtg ccttctagtt
     1621 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     1681 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     1741 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     1801 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     1861 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     1921 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     1981 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     2041 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     2101 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     2161 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     2221 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     2281 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     2341 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     2401 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     2461 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     2521 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     2581 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     2641 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg
     2701 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     2761 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     2821 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     2881 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     2941 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     3001 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     3061 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     3121 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     3181 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     3241 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     3301 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     3361 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     3421 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     3481 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     3541 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     3601 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     3661 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     3721 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3781 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     3841 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3901 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     3961 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     4021 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     4081 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     4141 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     4201 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     4261 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     4321 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4381 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4441 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4501 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4561 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4621 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4681 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4741 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tttttttgtt
     4801 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4861 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4921 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4981 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5041 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5101 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5161 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5221 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5281 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5341 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5401 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5461 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5521 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5581 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     5641 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5701 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5761 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5821 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5881 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5941 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6001 gacgtc
//
