LOCUS       sequence-230637-        5457 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  C-terminal tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5457)
  AUTHORS   Yesbolatova A, Natsume T, Hayashi KI, Kanemaki MT
  TITLE     Generation of conditional auxin-inducible degron (AID) cells and 
            tight control of degron-fused proteins using the degradation 
            inhibitor auxinole.
  JOURNAL   Methods. 2019 Apr 23. pii: S1046-2023(18)30331-1. doi: 
            10.1016/j.ymeth.2019.04.010.
  PUBMED    31026591
REFERENCE   2  (bases 1 to 5457)
  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: "Methods. 
            2019 Apr 23. pii: S1046-2023(18)30331-1. doi: 
            10.1016/j.ymeth.2019.04.010."
COMMENT     Sequence Label: pMK389 (mCherry-BSD)
FEATURES             Location/Qualifiers
     source          1..5457
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      4..459
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(91..110)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     301..322
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             683..1393
                     /codon_start=1
                     /product="derivative of the monomeric red fluorescent 
                     protein mCherry"
                     /label=mCherry2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFNWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQCRTMGWEASTERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVDIKLDILSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(828..846)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     polyA_signal    1511..1632
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1548..1567)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1602..1621
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1663..1696
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1757..2256
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1792..1815)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2189..2208
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      2280..2289
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2286..2684
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     protein_bind    3187..3220
                     /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(3269..3289)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3269..3287)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3308..3324)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3308..3324)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3321..3343)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3332..3348
                     /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(3356..3362,3363..3380,3381..3386))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3401..3422
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3539..3556)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3710..4298)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3790..3809)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4469..5260,5261..5329))
                     /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     5092..5111
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5330..5434)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
       61 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
      121 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
      181 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
      241 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
      301 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
      361 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
      421 ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggagctcgg
      661 atccggtgca ggcgccgcta gcatggtgag caagggcgag gaggataaca tggccatcat
      721 caaggagttc atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga
      781 gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt
      841 gaccaagggt ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg
      901 ctccaaggcc tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc
      961 cgagggcttc aattgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac
     1021 ccaggactcc tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa
     1081 cttcccctcc gacggccccg taatgcagtg tcgtaccatg ggctgggagg cctccactga
     1141 gcggatgtac cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa
     1201 ggacggcggc cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca
     1261 gctgcccggc gcctacaacg tcgacatcaa gttggacatc ctttcccaca acgaggacta
     1321 caccatcgtg gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga
     1381 gctgtacaag taatctagat aactgatcat aatcagccat accacatttg tagaggtttt
     1441 acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat
     1501 tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     1561 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     1621 caatgtatct taacgcgtcg atcatattca ataaccctta atataacttc gtataatgta
     1681 tgctatacga agttattagg tctgaagagg agtttacgtc cagccaagct taggatctcg
     1741 acctcgaaat tctaccgggt aggggaggcg cttttcccaa ggcagtctgg agcatgcgct
     1801 ttagcagccc cgctgggcac ttggcgctac acaagtggcc tctggcctcg cacacattcc
     1861 acatccaccg gtaggcgcca accgactccg ttctttggtg gccccttcgc gccaccttct
     1921 actcctcccc tagtcaggaa gttccccccc gccccgcagc tcgcgtcgtg caggacgtga
     1981 caaatggaag tagcacgtct cactagtctc gtgcagatgg acagcaccgc tgagcaatgg
     2041 aagcgggtag gcctttgggg cagcggccaa tagcagcttt gctccttcgc tttctgggct
     2101 cagaggctgg gaaggggtgg gtccgggggc gggctcaggg gcgggctcag gggcggggcg
     2161 ggcgcccgaa ggtcctccgg aggcccggca ttctgcacgc ttcaaaagcg cacgtctgcc
     2221 gcgctgttct cctcttcctc atctccgggc ctttcgacct gcatccatct agatctcgag
     2281 ccaccatggc caagcctttg tctcaagaag aatccaccct cattgaaaga gcaacggcta
     2341 caatcaacag catccccatc tctgaagact acagcgtcgc cagcgcagct ctctctagcg
     2401 acggccgcat cttcactggt gtcaatgtat atcattttac tgggggacct tgtgcagaac
     2461 tcgtggtgct gggcactgct gctgctgcgg cagctggcaa cctgacttgt atcgtcgcga
     2521 tcggaaatga gaacaggggc atcttgagcc cctgcggacg gtgccgacag gtgcttctcg
     2581 atctgcatcc tgggatcaaa gccatagtga aggacagtga tggacagccg acggcagttg
     2641 ggattcgtga attgctgccc tctggttatg tgtgggaggg ctaatcgatg atatcagatc
     2701 cccgggatgc agaaattgat gatctattaa acaataaaga tgtccactaa aatggaagtt
     2761 tttcctgtca tactttgtta agaagggtga gaacagagta cctacatttt gaatggaagg
     2821 attggagcta cgggggtggg ggtggggtgg gattagataa atgcctgctc tttactgaag
     2881 gctctttact attgctttat gataatgttt catagttgga tatcataatt taaacaagca
     2941 aaaccaaatt aagggccagc tcattcctcc cactcatgat ctatagatct atagatctct
     3001 cgtgggatca ttgtttttct cttgattccc actttgtggt tctaagtact gtggtttcca
     3061 aatgtgtcag tttcatagcc tgaagaacga gatcagcagc ctctgttcca catacacttc
     3121 attctcagta ttgttttgcc aagttctaat tccatcagaa gctggtcgag atccggaacc
     3181 cttaatataa cttcgtataa tgtatgctat acgaagttat taggtccctc gaagaggttc
     3241 actaggatcc ggtacccagc ttttgttccc tttagtgagg gttaattgcg cgcttggcgt
     3301 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     3361 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     3421 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     3481 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     3541 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     3601 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     3661 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     3721 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     3781 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     3841 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     3901 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     3961 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     4021 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     4081 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     4141 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     4201 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     4261 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     4321 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     4381 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     4441 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     4501 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     4561 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     4621 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     4681 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     4741 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     4801 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     4861 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     4921 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     4981 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     5041 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     5101 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     5161 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     5221 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     5281 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     5341 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     5401 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccac
//
