LOCUS       sequence-156869-        6151 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  14x MS2 sense + 14x PP7 antisense with loxP-kanMX-loxP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6151)
  AUTHORS   Lenstra TL, Coulon A, Chow CC, Larson DR
  TITLE     Single-Molecule Imaging Reveals a Switch between Spurious and 
            Functional ncRNA Transcription.
  JOURNAL   Mol Cell. 2015 Nov 19;60(4):597-610. doi: 
            10.1016/j.molcel.2015.09.028. Epub 2015 Nov 5.
  PUBMED    26549684
REFERENCE   2  (bases 1 to 6151)
  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.molcel.2015.09.028"; journalName: "Mol Cell"; date: 
            "2015-11-19- 19"; volume: "60"; issue: "4"; pages: "597-610"
COMMENT     Sequence Label: pTL032
FEATURES             Location/Qualifiers
     source          1..6151
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(1..792,793..861))
                     /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     624..643
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(862..966)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      993..1448
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1080..1099)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1290..1311
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1574..1596
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1588..1605
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1589..1605
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1615..1634
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1615..1633
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     1666..1682
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1666..1682
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    3361..3394
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            3449..4805
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        3449..3792
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             3793..4602
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3860..3879)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      4608..4805
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    4868..4901
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(4952..4972)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4952..4970)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4991..5007)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4991..5007)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5004..5026)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5015..5031
                     /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(5039..5045,5046..5063,5064..5069))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5084..5105
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5222..5239)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5393..5981)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5473..5492)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
       61 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
      121 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
      181 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
      241 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
      301 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
      361 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
      421 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
      481 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
      541 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
      601 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
      661 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
      721 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
      781 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
      841 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
      901 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
      961 tccgcgcaca tttccccgaa aagtgccacc tgacgcgccc tgtagcggcg cattaagcgc
     1021 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     1081 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     1141 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     1201 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     1261 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     1321 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     1381 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
     1441 tacaatttcc attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc
     1501 tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta
     1561 acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtgagcg cgcgtaatac
     1621 gactcactat agggcgaatt ggagctccac cgcggtggcg gccgctctag aactagtgga
     1681 tcctacggta cttattgcca agaaagcacg agcatcagcc gtgcctccag gtcgaatctt
     1741 caaacgacga cgatcacgcg tcgctccagt attccagggt tcatcagatc ctacggtact
     1801 tattgccaag aaagcacgag catcagccgt gcctccaggt cgaatcttca aacgacgacg
     1861 atcacgcgtc gctccagtat tccagggttc atcagatcct acggtactta ttgccaagaa
     1921 agcacgagca tcagccgtgc ctccaggtcg aatcttcaaa cgacgacgat cacgcgtcgc
     1981 tccagtattc cagggttcat cagatcctac ggtacttatt gccaagaaag cacgagcatc
     2041 agccgtgcct ccaggtcgaa tcttcaaacg acgacgatca cgcgtcgctc cagtattcca
     2101 gggttcatca gatcctacgg tacttattgc caagaaagca cgagcatcag ccgtgcctcc
     2161 aggtcgaatc ttcaaacgac gacgatcacg cgtcgctcca gtattccagg gttcatcaga
     2221 tcctacggta cttattgcca agaaagcacg agcatcagcc gtgcctccag gtcgaatctt
     2281 caaacgacga cgatcacgcg tcgctccagt attccagggt tcatcagatc ctacggtact
     2341 tattgccaag aaagcacgag catcagccgt gcctccaggt cgaatcttca aacgacgacg
     2401 atcacgcgtc gctccagtat tccagggttc atcagatccc ccgttctcga gatctaatga
     2461 acccgggaat actgcagcca gcgagcccat atgctctgct ggtttctaga gtcgacctgc
     2521 agggagcgac gccatatcgt ctgctccttt ctaggcaatt aggtacctta ggatctaatg
     2581 aacccgggaa tactgcagcc agcgagccca tatgctctgc tggtttctag agtcgacctg
     2641 cagggagcga cgccatatcg tctgctcctt tctaggcaat taggtacctt aggatctaat
     2701 gaacccggga atactgcagc cagcgagccc atatgctctg ctggtttcta gagtcgacct
     2761 gcagggagcg acgccatatc gtctgctcct ttctaggcaa ttaggtacct taggatctaa
     2821 tgaacccggg aatactgcag ccagcgagcc catatgctct gctggtttct agagtcgacc
     2881 tgcagggagc gacgccatat cgtctgctcc tttctaggca attaggtacc ttaggatcta
     2941 atgaacccgg gaatactgca gccagcgagc ccatatgctc tgctggtttc tagagtcgac
     3001 ctgcagggag cgacgccata tcgtctgctc ctttctaggc aattaggtac cttaggatct
     3061 aatgaacccg ggaatactgc agccagcgag cccatatgct ctgctggttt ctagagtcga
     3121 cctgcaggga gcgacgccat atcgtctgct cctttctagg caattaggta ccttaggatc
     3181 taatgaaccc gggaatactg cagccagcga gcccatatgc tctgctggtt tctagagtcg
     3241 acctgcaggg agcgacgcca tatcgtctgc tcctttctag gcaattaggt accttaggat
     3301 ctcctcgctc cactttgtag ggctgcagga attgtacgct gcaggtcgac aacccttaat
     3361 ataacttcgt ataatgtatg ctatacgaag ttattaggtc tagagatctg tttagcttgc
     3421 ctcgtccccg ccgggtcacc cggccagcga catggaggcc cagaataccc tccttgacag
     3481 tcttgacgtg cgcagctcag gggcatgatg tgactgtcgc ccgtacattt agcccataca
     3541 tccccatgta taatcatttg catccataca ttttgatggc cgcacggcgc gaagcaaaaa
     3601 ttacggctcc tcgctgcaga cctgcgagca gggaaacgct cccctcacag acgcgttgaa
     3661 ttgtccccac gccgcgcccc tgtagagaaa tataaaaggt taggatttgc cactgaggtt
     3721 cttctttcat atacttcctt ttaaaatctt gctaggatac agttctcaca tcacatccga
     3781 acataaacaa ccatgggtaa ggaaaagact cacgtttcga ggccgcgatt aaattccaac
     3841 atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca atcaggtgcg
     3901 acaatctatc gattgtatgg gaagcccgat gcgccagagt tgtttctgaa acatggcaaa
     3961 ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct gacggaattt
     4021 atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg gttactcacc
     4081 actgcgatcc ccggcaaaac agcattccag gtattagaag aatatcctga ttcaggtgaa
     4141 aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc tgtttgtaat
     4201 tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac
     4261 ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc
     4321 tggaaagaaa tgcataagct tttgccattc tcaccggatt cagtcgtcac tcatggtgat
     4381 ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat tgatgttgga
     4441 cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg cctcggtgag
     4501 ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa tcctgatatg
     4561 aataaattgc agtttcattt gatgctcgat gagtttttct aatcagtact gacaataaaa
     4621 agattcttgt tttcaagaac ttgtcatttg tatagttttt ttatattgta gttgttctat
     4681 tttaatcaaa tgttagcgtg atttatattt tttttcgcct cgacatcatc tgcccagatg
     4741 cgaagttaag tgcgcagaaa gtaatatcat gcgtcaatcg tatgtgaatg ctggtcgcta
     4801 tactgctgtc gattcgatac taacgccgcc atccagtgtc gaaaacgagc tctcgagaac
     4861 ccttaatata acttcgtata atgtatgcta tacgaagtta ttaggtgata tcagatccac
     4921 tagtggccta tgcggtaccc agcttttgtt ccctttagtg agggttaatt gcgcgcttgg
     4981 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     5041 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     5101 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     5161 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     5221 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     5281 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     5341 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     5401 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5461 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     5521 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     5581 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     5641 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     5701 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     5761 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     5821 gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     5881 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     5941 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     6001 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     6061 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     6121 aaagtatata tgagtaaact tggtctgaca g
//
