LOCUS       sequence-134603-        5951 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Floxed Kanamycin (G418) selection maker; 3' tagging vector used to 
            add 1x-mCherry followed by 12 repetitive PP7 version 4 RNA 
            stem-loops to gene of interest.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5951)
  AUTHORS   Saroufim MA, Bensidoun P, Raymond P, Rahman S, Krause MR, Oeffinger 
            M, Zenklusen D
  TITLE     The nuclear basket mediates perinuclear mRNA scanning in budding 
            yeast.
  JOURNAL   J Cell Biol. 2015 Dec 21;211(6):1131-40. doi: 10.1083/jcb.201503070.
  PUBMED    26694838
REFERENCE   2  (bases 1 to 5951)
  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: "10.1083/jcb"; 
            journalName: "J Cell Biol"; date: "2015-12-21- 21"; volume: "211"; 
            issue: "6"; pages: "1131-40"
COMMENT     Sequence Label: pDZ645 pKAN 1x-mCherry-12xPP7 V4
FEATURES             Location/Qualifiers
     source          1..5951
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     32..48
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     32..48
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     regulatory      54..63
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             60..770
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(205..223)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     473..492
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     777..794
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    1299..1332
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      1358..1545
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1413..1437)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1615..2971
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1615..1958
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1959..2768
                     /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(2026..2045)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2774..2971
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    3034..3067
                     /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(3118..3138)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3118..3136)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3157..3173)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3157..3173)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3170..3192)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3181..3197
                     /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(3205..3211,3212..3229,3230..3235))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3250..3271
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3388..3405)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3559..4147)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3639..3658)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4318..5109,5110..5178))
                     /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     4941..4960
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5179..5283)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5310..5765
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5397..5416)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5607..5628
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5891..5913
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5905..5922
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5906..5922
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5932..5951
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5932..5950
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 cgaattggag ctccaccgcg gtggcggccg ctctagaact agtggatccg gtcgccacca
       61 tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg cgcttcaagg
      121 tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag ggcgagggcc
      181 gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc cccctgccct
      241 tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac gtgaagcacc
      301 ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag tgggagcgcg
      361 tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg
      421 gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac ggccccgtaa
      481 tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc gaggacggcg
      541 ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac tacgacgctg
      601 aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca
      661 acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa cagtacgaac
      721 gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtaa catacgattt
      781 aggtgacact atagttaatt aaagatccat gcagaaccag cagagcatat gggctcgctg
      841 gctgcacgtc gactctagaa accagcagag catatgggct cgctggatgc agaaccagca
      901 gagcatatgg gctcgctggc tgcacgtcga ctctagaaac cagcagagca tatgggctcg
      961 ctggatgcag aaccagcaga gcatatgggc tcgctggctg cacgtcgact ctagaaacca
     1021 gcagagcata tgggctcgct ggatgcagaa ccagcagagc atatgggctc gctggctgca
     1081 cgtcgactct agaaaccagc agagcatatg ggctcgctgg atgcagaacc agcagagcat
     1141 atgggctcgc tggctgcacg tcgactctag aaaccagcag agcatatggg ctcgctggat
     1201 gcagaaccag cagagcatat gggctcgctg gctgcacgtc gactctagaa accagcagag
     1261 catatgggct cgctggatgc atagatctaa cccttaatat aacttcgtat aatgtatgct
     1321 atacgaagtt attaggtgat ctgccacttc taaataagcg aatttcttat gatttatgat
     1381 ttttattatt aaataagtta taaaaaaaat aagtgtatac aaattttaaa gtgactctta
     1441 ggttttaaaa cgaaaattct tattcttgag taactctttc ctgtaggtca ggttgctttc
     1501 tcaggtatag tatgaggtcg ctcttattga ccacacctct accggcagat ccgctaggga
     1561 taacagggtg gatctgttta gcttgcctcg tccccgccgg gtcacccggc cagcgacatg
     1621 gaggcccaga ataccctcct tgacagtctt gacgtgcgca gctcaggggc atgatgtgac
     1681 tgtcgcccgt acatttagcc catacatccc catgtataat catttgcatc catacatttt
     1741 gatggccgca cggcgcgaag caaaaattac ggctcctcgc tgcagacctg cgagcaggga
     1801 aacgctcccc tcacagacgc gttgaattgt ccccacgccg cgcccctgta gagaaatata
     1861 aaaggttagg atttgccact gaggttcttc tttcatatac ttccttttaa aatcttgcta
     1921 ggatacagtt ctcacatcac atccgaacat aaacaaccat gggtaaggaa aagactcacg
     1981 tttcgaggcc gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc
     2041 gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt gtatgggaag cccgatgcgc
     2101 cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg
     2161 tcagactaaa ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta
     2221 ctcctgatga tgcatggtta ctcaccactg cgatccccgg caaaacagca ttccaggtat
     2281 tagaagaata tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc
     2341 ggttgcattc gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg
     2401 ctcaggcgca atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc
     2461 gtaatggctg gcctgttgaa caagtctgga aagaaatgca taagcttttg ccattctcac
     2521 cggattcagt cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga
     2581 aattaatagg ttgtattgat gttggacgag tcggaatcgc agaccgatac caggatcttg
     2641 ccatcctatg gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa
     2701 aatatggtat tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt
     2761 ttttctaatc agtactgaca ataaaaagat tcttgttttc aagaacttgt catttgtata
     2821 gtttttttat attgtagttg ttctatttta atcaaatgtt agcgtgattt atattttttt
     2881 tcgcctcgac atcatctgcc cagatgcgaa gttaagtgcg cagaaagtaa tatcatgcgt
     2941 caatcgtatg tgaatgctgg tcgctatact gctgtcgatt cgatactaac gccgccatcc
     3001 agtgtcgaaa acgagctctc gagaaccctt aatataactt cgtataatgt atgctatacg
     3061 aagttattag gtgatatcag atccactagt ggcctatgcg gtacccagct tttgttccct
     3121 ttagtgaggg ttaattgcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     3181 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     3241 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     3301 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     3361 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     3421 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     3481 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     3541 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     3601 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     3661 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     3721 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     3781 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     3841 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     3901 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     3961 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     4021 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     4081 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     4141 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     4201 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     4261 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     4321 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     4381 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     4441 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     4501 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     4561 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     4621 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     4681 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     4741 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     4801 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     4861 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     4921 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     4981 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     5041 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     5101 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     5161 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     5221 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     5281 gcgcacattt ccccgaaaag tgccacctga cgcgccctgt agcggcgcat taagcgcggc
     5341 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     5401 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     5461 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     5521 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     5581 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     5641 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     5701 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac
     5761 aatttccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct
     5821 tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg
     5881 ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgagcgcgc gtaatacgac
     5941 tcactatagg g
//
