LOCUS       sequence-485522-        7302 bp    DNA     circular SYN 08-SEP-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7302)
  AUTHORS   Stewart DP, Koss B, Bathina M, Perciavalle RM, Bisanz K, Opferman JT
  TITLE     Ubiquitin-independent degradation of antiapoptotic MCL-1.
  JOURNAL   Mol Cell Biol. 2010 Jun;30(12):3099-110. Epub 2010 Apr 12.
  PUBMED    20385764
REFERENCE   2  (bases 1 to 7302)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Cell 
            Biol."; date: "2010-06"; volume: "30(12)"; pages: "3099-110. Epub 
            2010 Apr 12"
COMMENT     Sequence Label: p3XFlag-CMV10-Flag-mMcl-1
FEATURES             Location/Qualifiers
     source          1..7302
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     126..148
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     140..157
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     141..157
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        318..697
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        698..901
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     852..872
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             931..996
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             2001..2012
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2061..2683
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2260..2279)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2707..2727)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2725..3022
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             3076..3870
                     /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(3130..3149)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3740..3759
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4522..4656
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4559..4578)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4613..4632
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4661..4680)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4662..4680)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4694..4710)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4694..4710)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4707..4729)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4718..4734
                     /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(4742..4748,4749..4766,4767..4772))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4787..4808
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4925..4942)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5096..5684)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5176..5195)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5855..6646,6647..6715))
                     /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     6478..6497
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6716..6820)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6847..7275
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6934..6953)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7144..7165
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
       61 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
      121 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctgatct atacattgaa
      181 tcaatattgg caattagcca tattagtcat tggttatata gcataaatca atattggcta
      241 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      301 aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat caattacggg
      361 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      421 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      481 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      541 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      601 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      661 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      721 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      781 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      841 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      901 tcgtttagtg aaccgtcaga attaaccatg gactacaaag accatgacgg tgattataaa
      961 gatcatgaca tcgattacaa ggatgacgat gacaagcttg cggccgccat gtttggcctg
     1021 cggagaaacg cggtcatcgg cttgaacctg tactgcggcg gcgccagcct cggcgcgggc
     1081 ggcggttctc cggcaggggc gcgcctggtg gccgaggagg ccaaggcgcg gcgcgagggg
     1141 ggaggggagg ccgccctgct gcccggcgcg cgggtggtcg cccggccgcc gcccgtgggc
     1201 gccgaggacc ccgacgtcac cgcgtcggcc gaaaggcggc tgcataagtc gcccggcctc
     1261 ctcgccgtgc cgcccgagga gatggccgcg tcggccgccg ccgccatcgt gtctccggag
     1321 gaggaactgg acggctgcga gccggaggcc atcggcaagc gcccggccgt gctgcccctc
     1381 ctggagcgcg tgagcgaggc ggccaagagc tccggggccg acggctctct gccctccacg
     1441 ccgccgccgc ccgaggagga agaggacgac ctataccgcc agtcgctgga gatcatctcg
     1501 cgctacttgc gggagcaggc gaccggctcc aaggactcga agcctctggg cgaggcgggc
     1561 gcggcgggcc ggagagcgct ggagaccctg cggcgcgtgg gcgacggcgt gcagcgcaac
     1621 cacgagacgg ccttccaggg catgctccgg aaactggaca ttaaaaacga aggcgatgtt
     1681 aaatcttttt ctcgagtaat ggtccatgtt ttcaaagatg gcgtaacaaa ctggggcagg
     1741 attgtgactc ttatttcttt cggtgccttt gtggccaaac acttaaagag cgtaaaccaa
     1801 gaaagcttca tcgaaccatt agcagaaact atcacagatg ttcttgtaag gacgaaacgg
     1861 gactggcttg tcaaacaaag aggctgggat gggtttgtgg agttcttcca cgtacaggac
     1921 ctagaaggcg gcatcagaaa tgtgctgctg gcttttgcgg gtgttgctgg agtaggggct
     1981 ggtctggcat atctaataag atagaagggc gaattcatcg atagatctga tatcggtacc
     2041 agtcgactct agaggatccc gggtggcatc cctgtgaccc ctccccagtg cctctcctgg
     2101 ccctggaagt tgccactcca gtgcccacca gccttgtcct aataaaatta agttgcatca
     2161 ttttgtctga ctaggtgtcc ttctataata ttatggggtg gaggggggtg gtatggagca
     2221 aggggcaagt tgggaagaca acctgtaggg cctgcggggt ctattgggaa ccaagctgga
     2281 gtgcagtggc acaatcttgg ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc
     2341 tgcctcagcc tcccgagttg ttgggattcc aggcatgcat gaccaggctc agctaatttt
     2401 tgtttttttg gtagagacgg ggtttcacca tattggccag gctggtctcc aactcctaat
     2461 ctcaggtgat ctacccacct tggcctccca aattgctggg attacaggcg tgaaccactg
     2521 ctcccttccc tgtccttctg attttaaaat aactatacca gcaggaggac gtccagacac
     2581 agcataggct acctggccat gcccaaccgg tgggacattt gagttgcttg cttggcactg
     2641 tcctctcatg cgttgggtcc actcagtaga tgcctgttga attgggtacg cggccagctt
     2701 ggctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     2761 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     2821 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     2881 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     2941 gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga
     3001 agtagtgagg aggctttttt ggaggaattg atcagcttgg gatctgatca agagacagga
     3061 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     3121 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     3181 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     3241 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     3301 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     3361 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     3421 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     3481 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     3541 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     3601 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     3661 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     3721 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     3781 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     3841 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     3901 aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt
     3961 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     4021 tgctggagtt cttcgcccac cccgggctcg atcccctcgc gagttggttc agctgctgcc
     4081 tgaggctgga cgacctcgcg gagttctacc ggcagtgcaa atccgtcggc atccaggaaa
     4141 ccagcagcgg ctatccgcgc atccatgccc ccgaactgca ggagtgggga ggcacgatgg
     4201 ccgctttggt cgacccggac gggacgctcc tgcgcctgat acagaacgaa ttgcttgcag
     4261 gcatctcatg agtgtgtctt cccgttttcc gcctgaggtc actgcgtgga tggagcgctg
     4321 gcgcctgctg cgcgacggcg agctgctcac cacccactcg ccaagctgga accgtaaaaa
     4381 ggccgcgttg ctggcgtttt tccataggct ccgccgatca taatcagcca taccacattt
     4441 gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa
     4501 atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc
     4561 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     4621 tccaaactca tcaatgtatc ttatcatgtc tggatcaatt ccctatagtg agtcgtatta
     4681 aattcgtaat catgtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     4741 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     4801 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     4861 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     4921 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4981 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     5041 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     5101 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5161 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5221 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5281 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5341 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5401 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5461 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5521 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5581 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5641 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5701 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5761 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5821 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5881 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5941 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     6001 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     6061 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6121 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6181 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6241 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6301 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6361 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6421 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6481 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6541 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6601 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6661 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     6721 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6781 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6841 cacctgacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     6901 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     6961 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     7021 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     7081 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     7141 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     7201 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     7261 aatttaacgc gaattttaac aaaatattaa cgcttacaat tt
//
