LOCUS       sequence-364206-        5085 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tier-1 vector encoding PPGK-controlled 2x (GGGGS)-linker 
            (PPGK-2xGS-pA)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5085)
  AUTHORS   Haellman V, Strittmatter T, Bertschi A, Stucheli P, Fussenegger M
  TITLE     A versatile plasmid architecture for mammalian synthetic biology 
            (VAMSyB).
  JOURNAL   Metab Eng. 2021 Jul;66:41-50. doi: 10.1016/j.ymben.2021.04.003. Epub
            2021 Apr 18.
  PUBMED    33857582
REFERENCE   2  (bases 1 to 5085)
  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"; journalName: "Metab 
            Eng."; date: "2021-07"; volume: "66:41-50. doi"; pages: " 
            10.1016/j.ymben.2021.04.003. Epub 2021 Apr 18"
COMMENT     Sequence Label: pTS2412-Tier1-PPGK-2xGS
FEATURES             Location/Qualifiers
     source          1..5085
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(170..758)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(250..269)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(929..1720,1721..1789))
                     /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     1552..1571
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1790..1894)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1968..2467
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2003..2026)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2400..2419
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     complement(2601..2618)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2607..2831
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2877..3305
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2964..2983)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3174..3195
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3314..3334)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3319..3648
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3499..3634
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3561..3580
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3715..4509
                     /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(3769..3788)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4379..4398
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4683..4804
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4720..4739)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4774..4793
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4853..4869)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4853..4869)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4866..4888)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4877..4893
                     /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(4901..4907,4908..4925,4926..4931))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4946..4967
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(join(5084..5085,1..16))
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
       61 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
      121 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
      181 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
      241 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
      301 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
      361 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
      421 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
      481 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
      541 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
      601 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
      661 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
      721 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
      781 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
      841 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
      901 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
      961 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     1021 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     1081 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     1141 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     1201 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     1261 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     1321 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     1381 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     1441 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     1501 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     1561 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     1621 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     1681 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     1741 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     1801 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     1861 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     1921 acgtcgacgg atcgggagat ctccacgcgt ggtaccctcg agctaccggg taggggaggc
     1981 gcttttccca aggcagtctg gagcatgcgc tttagcagcc ccgctgggca cttggcgcta
     2041 cacaagtggc ctctggcctc gcacacattc cacatccacc ggtaggcgcc aaccggctcc
     2101 gttctttggt ggccccttcg cgccaccttc tactcctccc ctagtcagga agttcccccc
     2161 cgccccgcag ctcgcgtcgt gcaggacgtg acaaatggaa gtagcacgtc tcagtagtct
     2221 cgtgcagatg gacagcaccg ctgagcaatg gaagcgggta ggcctttggg gcagcggcca
     2281 atagcagctt tgctccttcg ctttctgggc tcagaggctg ggaaggggtg ggtccggggg
     2341 cgggctcagg ggcgggctca ggggcggggc gggcgcccga aggtcctccg gaggcccggc
     2401 attctgcacg cttcaaaagc gcacgtctgc cgcgctgttc tcctcttcct catctccggg
     2461 cctttcgacc tccagcccga gctcggtata gggagaccga attcaccatg actagtggag
     2521 gcggaggttc cggagggggg ggttccgcta gcggatccac cggtgtctag aaagctttga
     2581 ggccggcctg cagggatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     2641 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     2701 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     2761 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     2821 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     2881 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2941 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     3001 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     3061 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     3121 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     3181 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     3241 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     3301 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     3361 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     3421 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3481 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     3541 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
     3601 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
     3661 gcttgtatat ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt
     3721 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
     3781 gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag
     3841 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac
     3901 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
     3961 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
     4021 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
     4081 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
     4141 cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat
     4201 caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag
     4261 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
     4321 ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg
     4381 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
     4441 ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
     4501 ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat
     4561 cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc
     4621 gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc
     4681 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     4741 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     4801 cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc
     4861 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     4921 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     4981 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     5041 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcct
//
