LOCUS       sequence-110836-        8062 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  encodes the Int2 gene controlled by PBad promoter and a reversed 
            Int5 gene flanked by the attB/P sites of Int2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8062)
  AUTHORS   Yang L, Nielsen AA, Fernandez-Rodriguez J, McClune CJ, Laub MT, Lu 
            TK, Voigt CA
  TITLE     Permanent genetic memory with >1-byte capacity.
  JOURNAL   Nat Methods. 2014 Dec;11(12):1261-6. doi: 10.1038/nmeth.3147. Epub 
            2014 Oct 26.
  PUBMED    25344638
REFERENCE   2  (bases 1 to 8062)
  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: 
            "10.1038/nmeth.3147"; journalName: "Nat Methods"; date: "2014-12"; 
            volume: "11"; issue: "12"; pages: "1261-6"
COMMENT     Sequence Label: pCas_2+5
FEATURES             Location/Qualifiers
     source          1..8062
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        43..77
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     misc_RNA        complement(1856..1905)
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     terminator      complement(1977..2004)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(2020..2091)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(2536..2555)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2547..3362
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             2547..3362
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2620..2639)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3479..3496
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      4070..4658
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4559..4578
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(5140..5169)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     CDS             complement(5216..6094)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        6121..6405
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     6328..6347
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
ORIGIN
        1 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atttgacagc tagctcagtc
       61 ctaggtataa tgctagcagt tcgatgagag cgataacgga cggcgcagaa ggggagtagc
      121 tcttcgccgg accgtcgaca tactgctcag ctcgtcttag gtacgggctg cccatttgat
      181 tttaacgcgt tcatcaccat caaacggacg accacgctgg ccttttgcaa cccaaatttc
      241 atcgatgcag gtatcaataa ttgcattacg catggtcggg gttgcacgca gccacagttc
      301 ttcataatcg ctgctatcaa caatccagct aacatcaact gctgcgcttg cgctgctttc
      361 gctaactgca tctttggctg cctgcagggt gctcagtgct tcttgatatg caggggcaaa
      421 aaactgttct gccggaccat cataaacacc attctgacga tcacgcagca ggcgacccag
      481 atttttttcg gcttcacgaa ctgcggcttt tgcatacttt tcatcttcgc ttgcctgcgg
      541 attggtcagt gctgcccagc gatctgcaac tgcaataaca aacggatcat ccggttcgct
      601 tgctgctaat tttgctgccc aacgaaatgc aacatattct tcaacgcttt tacgtgcaac
      661 ataggtcggt gccggacaac cacctttcac actgctacgc caacaacgat aaccattacc
      721 gctatagcta cagctaccac cacaacccgg acaacgcata cgaccgctca gcagatgttt
      781 gcgacgggta tcatgatcgc taccatccag cggaacacca acaccatctt cacctttaac
      841 ggctgctttt gcggcttctt gttcttcatc ggtcaccagc ggaggaccat gcataacgct
      901 aacacgttta ccttcaccgt tataaaaggt cagacgacgc tgtttaccat cctgacgacc
      961 tgtggtctgc caacccgcat atgccggatt ctgaatcata tcacgcacgg taactgcaat
     1021 ccacggacca ccggtcgggc tcggaatttc acgggtattc attgcatgtg cggtgcctgc
     1081 atagctcaga cgatcggtaa ccggcagggt aaaaaccaga cgggctgctt ctgctttggt
     1141 cagaccatca ggaccacccg catcttcatc atctgctgcc agtttacgtt catcatattc
     1201 atcaccctct tcatcactaa cggtaaccag aacaacacgc agaccatacg gtgcacgggc
     1261 attaacccat tcaccatttt cacgctgatg tgctttggta tcacgaacac gttcgctcag
     1321 tttttctgct tcttcgcgtg cttcttctgc acgacgaatc agttcaccgc gatcacgttt
     1381 attggtgcta tccagaaccg gacgaccggt atcttcatcc caaccaaaca gcagacgacg
     1441 aggcatacca tcttccggtt cgataatttt cagaattgca ccggcaccac cacgatccca
     1501 acgatccaga cgataacacc acagtgcacc aacttcaccg ctttccaggg ctttcagtgc
     1561 tttgctctga tcatcacgtg ctttaccttt acgaaaacgg cttgcgctac caacttcttt
     1621 ccaaacatga cgaacctgca tacccagcag tgctgcaact ttacgaccca gggtttcttg
     1681 tgctgcaatg ctaatttctt gtttacgacg ctgacctgca ccatttgcac ggcttttaac
     1741 tgctttgctt ttacgacaaa acaggtcaat cagacctgca ggatccggac cggtttcggt
     1801 ggtcatacca ggcatcgagc ctccttcctc tgttaaacaa aattatttgt agaggctgtt
     1861 tcgtcctcac ggactcatca gaccggaaag cacatccggt gacagctagc caagagcagt
     1921 gccttgcaga agttctcggg cgagaatctc gcgtagacac atacatgagc tataaacgca
     1981 gaaaggccca cccgaaggtg agccagtgtg actctagtag agagcgttca ccgacaaaca
     2041 acagataaaa cgaaaggccc agtctttcga ctgagccttt cgttttattt gatgcctggc
     2101 tagggcccat acccaaaaaa aaaccccgcc cctgacaggg cggggttttt tttaggtaga
     2161 gacccacgag gcagacctca gcgctagcgc gaaaagtctt catatctttc ggctagcgct
     2221 gaggtctgcc tcgtgaagaa ggtgttgctg actcatacca ggcctgaatc gccccatcat
     2281 ccagccagaa agtgagggag ccacggttga tgagagcttt gttgtaggtg gaccagttgg
     2341 tgattttgaa cttttgcttt gccacggaac ggtctgcgtt gtcgggaaga tgcgtgatct
     2401 gatccttcaa ctcagcaaaa gttcgattta ttcaacaaag ccacgttgtg tctcaaaatc
     2461 tctgatgtta cattgcacaa gataaaaata tatcatcatg aacaataaaa ctgtctgctt
     2521 acataaacag taatacaagg ggtgttatga gccatattca acgggaaacg tcttgctcga
     2581 ggccgcgatt aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata
     2641 atgtcgggca atcaggtgcg acaatctatc gattgtatgg gaagcccgat gcgccagagt
     2701 tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac
     2761 taaactggct gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg
     2821 atgatgcatg gttactcacc actgcgatcc ccgggaaaac agcattccag gtattagaag
     2881 aatatcctga ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc
     2941 attcgattcc tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg
     3001 cgcaatcacg aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg
     3061 gctggcctgt tgaacaagtc tggaaagaaa tgcataagct tttgccattc tcaccggatt
     3121 cagtcgtcac tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa
     3181 taggttgtat tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc
     3241 tatggaactg cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg
     3301 gtattgataa tcctgatatg aataaattgc agtttcattt gatgctcgat gagtttttct
     3361 aatcagaatt ggttaattgg ttgtaacact ggcagagcat tacgctgact tgacgggact
     3421 cgagattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     3481 cgagtcagtg agcgaggaag cctgcataac gcgaagtaat cttttcggtt ttaaagaaaa
     3541 agggcagggt ggtgacacct tgcccgtttt tttgccggac tgcagcaatg gcaacaacgt
     3601 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     3661 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     3721 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     3781 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     3841 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     3901 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     3961 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     4021 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     4081 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     4141 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     4201 agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     4261 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     4321 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     4381 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     4441 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     4501 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     4561 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     4621 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     4681 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     4741 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     4801 cgaccgagcg cagcgaggtc ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg
     4861 atgccggcca cgatgcgtcc ggcgtagagg atctgctcat gtttgacagc ttatcatcga
     4921 tgcataatgt gcctgtcaaa tggacgaagc agggattctg caaaccctat gctactccct
     4981 cgagccgtca attgtctgat tcgttaccaa ttatactagc tccggcaaaa aaacgggcaa
     5041 ggtgtcacca ccctgccctt tttctttaaa accgaaaaga ttacttcgcg tttgccacct
     5101 gacgtctaag aaaaggaata ttcagcaatt tgcccgtgcc gaagaaaggc ccacccgtga
     5161 aggtgagcca gtgagttgat tgctacgtaa ttagttagtt agcccttagt gactcttatg
     5221 acaacttgac ggctacatca ttcacttttt cttcacaacc ggcacggaac tcgctcgggc
     5281 tggccccggt gcatttttta aatacccgcg agaaatagag ttgatcgtca aaaccaacat
     5341 tgcgaccgac ggtggcgata ggcatccggg tggtgctcaa aagcagcttc gcctggctga
     5401 tacgttggtc ctcgcgccag cttaagacgc taatccctaa ctgctggcgg aaaagatgtg
     5461 acagacgcga cggcgacaag caaacatgct gtgcgacgct ggcgatatca aaattgctgt
     5521 ctgccaggtg atcgctgatg tactgacaag cctcgcgtac ccgattatcc atcggtggat
     5581 ggagcgactc gttaatcgct tccatgcgcc gcagtaacaa ttgctcaagc agatttatcg
     5641 ccagcagctc cgaatagcgc ccttcccctt gcccggcgtt aatgatttgc ccaaacaggt
     5701 cgctgaaatg cggctggtgc gcttcatccg ggcgaaagaa ccccgtattg gcaaatattg
     5761 acggccagtt aagccattca tgccagtagg cgcgcggacg aaagtaaacc cactggtgat
     5821 accattcgcg agcctccgga tgacgaccgt agtgatgaat ctctcctggc gggaacagca
     5881 aaatatcacc cggtcggcaa acaaattctc gtccctgatt tttcaccacc ccctgaccgc
     5941 gaatggtgag attgagaata taacctttca ttcccagcgg tcggtcgata aaaaaatcga
     6001 gataaccgtt ggcctcaatc ggcgttaaac ccgccaccag atgggcatta aacgagtatc
     6061 ccggcagcag gggatcattt tgcgcttcag ccatactttt catactcccg ccattcagag
     6121 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
     6181 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
     6241 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
     6301 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
     6361 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg tttttttggg
     6421 ctagcaacat agcgaatcgt aaggagttaa aagatgccga ttgcaccgga atttctgagc
     6481 ctggcatatc cgggtcaaga atttccggca tatctgtatg gtcgtgcaag ccgtgatccg
     6541 aaacgtaaag gtcgtagcgt tcagagccag ctggatgaag gtcgtgccac ctgtctggat
     6601 gcaggttggc cgattgccgg tgaatttaaa gatgttgatc gtagcgcaag cgcatatgca
     6661 cgtcgtaccc gtgatgaatt tgaagaaatg attgcaggta ttcaggcagg cgaatgtcgt
     6721 attctggttg catttgaagc aagccgttat tatcgtgatc tggaagcata tgttcgtctg
     6781 cgtcgtgttt gtcgtgaagc cggtgttctg ctgtgttata atggtcaggt ttatgatctg
     6841 agcaaaagcg cagatcgtaa agcaaccgca caggatgcag ttaatgccga aggtgaagca
     6901 gatgatattc gtgaacgtaa tctgcgtacc acccgtctga atgcaaaacg tggtggtgca
     6961 catggtccgg ttccggatgg ttataaacgt cgttatgatc cggatagcgg tgatctggtt
     7021 gatcagattc cgcatcctga tcgtgcaggt ctgattaccg aaatttttcg tcgtgcagca
     7081 gcagccgaac cgctggcagc aatttgccgt gatctgaatg aacgtggtga aaccacccat
     7141 cgtggtaaag catggcagcg tcatcatctg catgcaattc tgcgtaatcc tgcatatatt
     7201 ggtcatcgtc gtcatctggg tgttgatacc ggtaaaggta tgtgggcacc gatttgtgat
     7261 gatgaagatt ttgcagaaac ctttcaggcc gttcaagaaa ttctgagtct gcctggtcgt
     7321 cagctgagtc cgggtccgga agcacagcat ctgcagaccg gtattgcact gtgtggtgaa
     7381 catcctgatg aaccgcctct gcgtagcgtt accgttcgtg gtcgtaccaa ttataactgt
     7441 agcacccgtt atgatgttgc catgcgtgaa gatcgtatgg atgcatttgt tgaagaaagc
     7501 gtgattacct ggctggcaag tgatgaagca gttgcagcct ttgaagataa caccgatgat
     7561 gaacgtaccc gtaaagcacg tattcgtctg aaagttctgg aagaacagct ggaagcagca
     7621 cagaaacagg cacgtaccct gcgtccggat ggcatgggta tgctgctgag cattgatagc
     7681 ctggcaggtc tggaagccga actgacaccg cagattgata aagcccgtca agaaagccgt
     7741 agcctgcatg ttccggcact gctgcgtgat ctgctgggta aaccgcgtgc agatgtggat
     7801 cgtgcatgga atgaagcact gaccctgccg cagcgtcgca tgattctgcg catggttgtt
     7861 accattcgtc tgtttaaagc aggtagccgt ggtgttcgtg caattgaacc gggtcgtatt
     7921 accctgagct atgttggcga accgggtttt aaaccggttg gtggtaatcg tgcaaaacag
     7981 taacagataa aaaaaatcct tagctttcgc taaggatgat ttctgattag ggtgatggtt
     8041 cacgtagtgg gccatcgact tg
//
