LOCUS       Exported                8939 bp ds-DNA     circular SYN 10-DEC-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8939)
  AUTHORS   Noboru Mizushima
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Jan 28, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8939
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          1852..5121
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     60..82
                     /label=AA_674
     promoter        386..661
                     /label=chicken beta-actin promoter
     primer_bind     546..557
                     /label=AA_628
     primer_bind     590..612
                     /label=AA_675
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1149..1171
                     /label=AA_676
     primer_bind     1677..1700
                     /label=AA_819
     primer_bind     1678..1700
                     /label=AA_677
     CDS             1741..1806
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    1816..1840
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(1821..1875)
                     /label=AA_666
     primer_bind     1829..1877
                     /label=AA_664
     primer_bind     complement(1829..1851)
                     /label=AA_667
     misc_feature    1852..5121
                     /label=dPspCas13b H133A/H1058A
     primer_bind     1852..1877
                     /label=AA_665
     primer_bind     complement(1908..1930)
                     /label=AA_684
     misc_feature    2233..2250
                     /label=HEPN1
     primer_bind     2490..2514
                     /label=AA_020F
                     /note="dPspcas13b primer F1"
     primer_bind     3133..3157
                     /label=AA_021F
                     /note="dPspCas13b primer F2"
     primer_bind     3793..3817
                     /label=AA_022F
                     /note="dPspCas13b primerF3"
     misc_feature    5008..5025
                     /label=HEPN2
     primer_bind     5038..5057
                     /label=AA_313
     primer_bind     complement(5082..5144)
                     /label=AA_673
     primer_bind     5099..5151
                     /label=AA_672
     primer_bind     5122..5151
                     /label=AA_671
     CDS             5131..5157
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             5167..5874
                     /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     5167..5189
                     /label=AA_628
     primer_bind     5438..5457
                     /label=mScarlet-colonyPCR-F
     primer_bind     5787..5807
                     /label=08 pcold_mcherry_seq-F
     primer_bind     complement(5848..5897)
                     /label=AA_668
     primer_bind     complement(5855..5874)
                     /label=EGFP-R
     primer_bind     complement(5855..5874)
                     /label=EGFP-Ra
     primer_bind     complement(5855..5874)
                     /label=YSPB008_GFP_R
     primer_bind     5861..5897
                     /label=AA_669
     primer_bind     5872..5901
                     /label=AA_670
     CDS             5875..5877
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation=""
     primer_bind     complement(5933..5957)
                     /label=AA_820
     polyA_site      6028..6083
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal"
     primer_bind     6396..6418
                     /label=AA_678
     primer_bind     complement(6442..6458)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6466..6482
                     /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(6490..6520)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6535..6556
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6615..6810
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      6661..6796
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    6816..6950
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     6947..6969
                     /label=AA_679
     rep_origin      complement(7189..7777)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7487..7509
                     /label=AA_680
     CDS             complement(7948..8808)
                     /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     8076..8098
                     /label=AA_681
     primer_bind     8683..8705
                     /label=AA_682
     promoter        complement(8809..8913)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attcctcgag gatcaccatg
     1741 gactacaaag accatgacgg tgattataaa gatcatgaca tcgattacaa ggatgacgat
     1801 gacaagcacc ggtatacaag tttgtacaaa aaagcaggct ccgcggccgc tatgaacatc
     1861 cccgctctgg tggaaaacca gaagaagtac tttggcacct acagcgtgat ggccatgctg
     1921 aacgctcaga ccgtgctgga ccacatccag aaggtggccg atattgaggg cgagcagaac
     1981 gagaacaacg agaatctgtg gtttcacccc gtgatgagcc acctgtacaa cgccaagaac
     2041 ggctacgaca agcagcccga gaaaaccatg ttcatcatcg agcggctgca gagctacttc
     2101 ccattcctga agatcatggc cgagaaccag agagagtaca gcaacggcaa gtacaagcag
     2161 aaccgcgtgg aagtgaacag caacgacatc ttcgaggtgc tgaagcgcgc cttcggcgtg
     2221 ctgaagatgt acagggacct gaccaacgca tacaagacct acgaggaaaa gctgaacgac
     2281 ggctgcgagt tcctgaccag cacagagcaa cctctgagcg gcatgatcaa caactactac
     2341 acagtggccc tgcggaacat gaacgagaga tacggctaca agacagagga cctggccttc
     2401 atccaggaca agcggttcaa gttcgtgaag gacgcctacg gcaagaaaaa gtcccaagtg
     2461 aataccggat tcttcctgag cctgcaggac tacaacggcg acacacagaa gaagctgcac
     2521 ctgagcggag tgggaatcgc cctgctgatc tgcctgttcc tggacaagca gtacatcaac
     2581 atctttctga gcaggctgcc catcttctcc agctacaatg cccagagcga ggaacggcgg
     2641 atcatcatca gatccttcgg catcaacagc atcaagctgc ccaaggaccg gatccacagc
     2701 gagaagtcca acaagagcgt ggccatggat atgctcaacg aagtgaagcg gtgccccgac
     2761 gagctgttca caacactgtc tgccgagaag cagtcccggt tcagaatcat cagcgacgac
     2821 cacaatgaag tgctgatgaa gcggagcagc gacagattcg tgcctctgct gctgcagtat
     2881 atcgattacg gcaagctgtt cgaccacatc aggttccacg tgaacatggg caagctgaga
     2941 tacctgctga aggccgacaa gacctgcatc gacggccaga ccagagtcag agtgatcgag
     3001 cagcccctga acggcttcgg cagactggaa gaggccgaga caatgcggaa gcaagagaac
     3061 ggcaccttcg gcaacagcgg catccggatc agagacttcg agaacatgaa gcgggacgac
     3121 gccaatcctg ccaactatcc ctacatcgtg gacacctaca cacactacat cctggaaaac
     3181 aacaaggtcg agatgtttat caacgacaaa gaggacagcg ccccactgct gcccgtgatc
     3241 gaggatgata gatacgtggt caagacaatc cccagctgcc ggatgagcac cctggaaatt
     3301 ccagccatgg ccttccacat gtttctgttc ggcagcaaga aaaccgagaa gctgatcgtg
     3361 gacgtgcaca accggtacaa gagactgttc caggccatgc agaaagaaga agtgaccgcc
     3421 gagaatatcg ccagcttcgg aatcgccgag agcgacctgc ctcagaagat cctggatctg
     3481 atcagcggca atgcccacgg caaggatgtg gacgccttca tcagactgac cgtggacgac
     3541 atgctgaccg acaccgagcg gagaatcaag agattcaagg acgaccggaa gtccattcgg
     3601 agcgccgaca acaagatggg aaagagaggc ttcaagcaga tctccacagg caagctggcc
     3661 gacttcctgg ccaaggacat cgtgctgttt cagcccagcg tgaacgatgg cgagaacaag
     3721 atcaccggcc tgaactaccg gatcatgcag agcgccattg ccgtgtacga tagcggcgac
     3781 gattacgagg ccaagcagca gttcaagctg atgttcgaga aggcccggct gatcggcaag
     3841 ggcacaacag agcctcatcc atttctgtac aaggtgttcg cccgcagcat ccccgccaat
     3901 gccgtcgagt tctacgagcg ctacctgatc gagcggaagt tctacctgac cggcctgtcc
     3961 aacgagatca agaaaggcaa cagagtggat gtgcccttca tccggcggga ccagaacaag
     4021 tggaaaacac ccgccatgaa gaccctgggc agaatctaca gcgaggatct gcccgtggaa
     4081 ctgcccagac agatgttcga caatgagatc aagtcccacc tgaagtccct gccacagatg
     4141 gaaggcatcg acttcaacaa tgccaacgtg acctatctga tcgccgagta catgaagaga
     4201 gtgctggacg acgacttcca gaccttctac cagtggaacc gcaactaccg gtacatggac
     4261 atgcttaagg gcgagtacga cagaaagggc tccctgcagc actgcttcac cagcgtggaa
     4321 gagagagaag gcctctggaa agagcgggcc tccagaacag agcggtacag aaagcaggcc
     4381 agcaacaaga tccgcagcaa ccggcagatg agaaacgcca gcagcgaaga gatcgagaca
     4441 atcctggata agcggctgag caacagccgg aacgagtacc agaaaagcga gaaagtgatc
     4501 cggcgctaca gagtgcagga tgccctgctg tttctgctgg ccaaaaagac cctgaccgaa
     4561 ctggccgatt tcgacggcga gaggttcaaa ctgaaagaaa tcatgcccga cgccgagaag
     4621 ggaatcctga gcgagatcat gcccatgagc ttcaccttcg agaaaggcgg caagaagtac
     4681 accatcacca gcgagggcat gaagctgaag aactacggcg acttctttgt gctggctagc
     4741 gacaagagga tcggcaacct gctggaactc gtgggcagcg acatcgtgtc caaagaggat
     4801 atcatggaag agttcaacaa atacgaccag tgcaggcccg agatcagctc catcgtgttc
     4861 aacctggaaa agtgggcctt cgacacatac cccgagctgt ctgccagagt ggaccgggaa
     4921 gagaaggtgg acttcaagag catcctgaaa atcctgctga acaacaagaa catcaacaaa
     4981 gagcagagcg acatcctgcg gaagatccgg aacgccttcg atgcaaacaa ttaccccgac
     5041 aaaggcgtgg tggaaatcaa ggccctgcct gagatcgcca tgagcatcaa gaaggccttt
     5101 ggggagtacg ccatcatgaa gggatccgga ctgcctccac ttgaaagact gacactggcg
     5161 gccgctatgg tgagcaaggg cgaggaggat aacatggcca tcatcaagga gttcatgcgc
     5221 ttcaaggtgc acatggaggg ctccgtgaac ggccacgagt tcgagatcga gggcgagggc
     5281 gagggccgcc cctacgaggg cacccagacc gccaagctga aggtgaccaa gggtggcccc
     5341 ctgcccttcg cctgggacat cctgtcccct cagttcatgt acggctccaa ggcctacgtg
     5401 aagcaccccg ccgacatccc cgactacttg aagctgtcct tccccgaggg cttcaagtgg
     5461 gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg tgacccagga ctcctccctg
     5521 caggacggcg agttcatcta caaggtgaag ctgcgcggca ccaacttccc ctccgacggc
     5581 cccgtaatgc agaagaagac catgggctgg gaggcctcct ccgagcggat gtaccccgag
     5641 gacggcgccc tgaagggcga gatcaagcag aggctgaagc tgaaggacgg cggccactac
     5701 gacgctgagg tcaagaccac ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac
     5761 aacgtcaaca tcaagttgga catcacctcc cacaacgagg actacaccat cgtggaacag
     5821 tacgaacgcg ccgagggccg ccactccacc ggcggcatgg acgagctgta caagtaagcg
     5881 gccgcactcc tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc
     5941 tggctcacaa ataccactga gatctttttc cctctgccaa aaattatggg gacatcatga
     6001 agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt
     6061 gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttaaaac
     6121 atcagaatga gtatttggtt tagagtttgg caacatatgc catatgctgg ctgccatgaa
     6181 caaaggtggc tataaagagg tcatcagtat atgaaacagc cccctgctgt ccattcctta
     6241 ttccatagaa aagccttgac ttgaggttag atttttttta tattttgttt tgtgttattt
     6301 ttttctttaa catccctaaa attttcctta catgttttac tagccagatt tttcctcctc
     6361 tcctgactac tcccagtcat agctgtccct cttctcttat gaagatccct cgacctgcag
     6421 cccaagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     6481 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     6541 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     6601 gtgccagcgg atccgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     6661 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     6721 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     6781 ggcttttttg gaggcctagg cttttgcaaa aagctaactt gtttattgca gcttataatg
     6841 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     6901 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc cgctgcatta
     6961 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     7021 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     7081 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     7141 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     7201 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     7261 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     7321 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     7381 tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     7441 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     7501 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     7561 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     7621 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     7681 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     7741 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     7801 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     7861 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     7921 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     7981 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     8041 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     8101 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     8161 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     8221 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     8281 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     8341 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     8401 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     8461 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     8521 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     8581 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     8641 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     8701 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     8761 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     8821 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     8881 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctg
//
