LOCUS       sequence-234560-        5466 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 5466)
  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 5466)
  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: pMK388 (mClover-BSD)
FEATURES             Location/Qualifiers
     source          1..5466
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     42..64
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     56..73
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     57..73
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     83..102
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        83..101
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(140..142,143..145,146..859)
                     /codon_start=1
                     /product="bright green-yellow fluorescent protein derived 
                     from mClover2, with improved photostability (Bajar et al., 
                     2003)"
                     /label=mClover3
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(185..206)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     793..814
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    977..1098
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1014..1033)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1068..1087
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1129..1162
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1223..1722
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1258..1281)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1655..1674
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      1746..1755
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1752..2150
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     protein_bind    2653..2686
                     /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(2735..2755)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2735..2753)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2774..2790)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2774..2790)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2787..2809)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2798..2814
                     /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(2822..2828,2829..2846,2847..2852))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2867..2888
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3005..3022)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3176..3764)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3256..3275)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3935..4726,4727..4795))
                     /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     4558..4577
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4796..4900)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4927..5382
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5014..5033)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5224..5245
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
       61 aacgacggcc agtgagcgcg cgtaatacga ctcactatag ggcgaattgg agctcggatc
      121 cggtgcaggc gccgctagca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
      181 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtcc gcggcgaggg
      241 cgagggcgat gccaccaacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
      301 gcccgtgccc tggcccaccc tcgtgaccac cttcggctac ggcgtggcct gcttcagccg
      361 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
      421 ccaggagcgc accatctctt tcaaggacga cggtacctac aagacccgcg ccgaggtgaa
      481 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
      541 cggcaacatc ctggggcaca agctggagta caacttcaac agccacaacg tctatatcac
      601 ggccgacaag cagaagaacg gcatcaaggc taacttcaag atccgccaca acgttgagga
      661 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
      721 gctgctgccc gacaaccact acctgagcca tcagtccaag ctgagcaaag accccaacga
      781 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatta cacatggcat
      841 ggacgagctg tacaagtaat ctagataact gatcataatc agccatacca catttgtaga
      901 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
      961 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     1021 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     1081 actcatcaat gtatcttaac gcgtcgatca tattcaataa cccttaatat aacttcgtat
     1141 aatgtatgct atacgaagtt attaggtctg aagaggagtt tacgtccagc caagcttagg
     1201 atctcgacct cgaaattcta ccgggtaggg gaggcgcttt tcccaaggca gtctggagca
     1261 tgcgctttag cagccccgct gggcacttgg cgctacacaa gtggcctctg gcctcgcaca
     1321 cattccacat ccaccggtag gcgccaaccg actccgttct ttggtggccc cttcgcgcca
     1381 ccttctactc ctcccctagt caggaagttc ccccccgccc cgcagctcgc gtcgtgcagg
     1441 acgtgacaaa tggaagtagc acgtctcact agtctcgtgc agatggacag caccgctgag
     1501 caatggaagc gggtaggcct ttggggcagc ggccaatagc agctttgctc cttcgctttc
     1561 tgggctcaga ggctgggaag gggtgggtcc gggggcgggc tcaggggcgg gctcaggggc
     1621 ggggcgggcg cccgaaggtc ctccggaggc ccggcattct gcacgcttca aaagcgcacg
     1681 tctgccgcgc tgttctcctc ttcctcatct ccgggccttt cgacctgcat ccatctagat
     1741 ctcgagccac catggccaag cctttgtctc aagaagaatc caccctcatt gaaagagcaa
     1801 cggctacaat caacagcatc cccatctctg aagactacag cgtcgccagc gcagctctct
     1861 ctagcgacgg ccgcatcttc actggtgtca atgtatatca ttttactggg ggaccttgtg
     1921 cagaactcgt ggtgctgggc actgctgctg ctgcggcagc tggcaacctg acttgtatcg
     1981 tcgcgatcgg aaatgagaac aggggcatct tgagcccctg cggacggtgc cgacaggtgc
     2041 ttctcgatct gcatcctggg atcaaagcca tagtgaagga cagtgatgga cagccgacgg
     2101 cagttgggat tcgtgaattg ctgccctctg gttatgtgtg ggagggctaa tcgatgatat
     2161 cagatccccg ggatgcagaa attgatgatc tattaaacaa taaagatgtc cactaaaatg
     2221 gaagtttttc ctgtcatact ttgttaagaa gggtgagaac agagtaccta cattttgaat
     2281 ggaaggattg gagctacggg ggtgggggtg gggtgggatt agataaatgc ctgctcttta
     2341 ctgaaggctc tttactattg ctttatgata atgtttcata gttggatatc ataatttaaa
     2401 caagcaaaac caaattaagg gccagctcat tcctcccact catgatctat agatctatag
     2461 atctctcgtg ggatcattgt ttttctcttg attcccactt tgtggttcta agtactgtgg
     2521 tttccaaatg tgtcagtttc atagcctgaa gaacgagatc agcagcctct gttccacata
     2581 cacttcattc tcagtattgt tttgccaagt tctaattcca tcagaagctg gtcgagatcc
     2641 ggaaccctta atataacttc gtataatgta tgctatacga agttattagg tccctcgaag
     2701 aggttcacta ggatccggta cccagctttt gttcccttta gtgagggtta attgcgcgct
     2761 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     2821 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     2881 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     2941 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     3001 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     3061 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     3121 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     3181 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     3241 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     3301 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     3361 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     3421 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     3481 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     3541 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     3601 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     3661 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     3721 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     3781 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     3841 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     3901 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     3961 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     4021 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     4081 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     4141 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     4201 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     4261 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     4321 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     4381 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     4441 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     4501 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     4561 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     4621 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     4681 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     4741 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     4801 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     4861 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     4921 cacctgacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     4981 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     5041 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     5101 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     5161 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     5221 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     5281 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     5341 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttccattcgc cattcaggct
     5401 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
     5461 aggggg
//
