LOCUS       sequence-402768-        4743 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Ec86 ncRNA in Escherichia coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4743)
  AUTHORS   Wang Y, Guan Z, Wang C, Nie Y, Chen Y, Qian Z, Cui Y, Xu H, Wang Q, 
            Zhao F, Zhang D, Tao P, Sun M, Yin P, Jin S, Wu S, Zou T
  TITLE     Cryo-EM structures of Escherichia coli Ec86 retron complexes reveal 
            architecture and defence mechanism.
  JOURNAL   Nat Microbiol. 2022 Sep;7(9):1480-1489. doi: 
            10.1038/s41564-022-01197-7. Epub 2022 Aug 18.
  PUBMED    35982312
REFERENCE   2  (bases 1 to 4743)
  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/s41564-022-01197-7"; journalName: "Nat Microbiol"; date: 
            "2022-09"; volume: "7"; issue: "9"; pages: "1480-1489"
COMMENT     Sequence Label: pBB75-ncRNA
FEATURES             Location/Qualifiers
     source          1..4743
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1037..1042,1043..1060,1061..1067)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1075..1091
                     /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)."
     primer_bind     1080..1102
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1099..1115
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1099..1115
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1111..1641
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVISIPRN"
     primer_bind     complement(1131..1148)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1131..1147)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1140..1162)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1639..1658
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1639..1657
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             1689..1711
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(1987..2005)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2001..2048
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(2119..2137)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      2684..3229
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     3130..3149
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3346..3363
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3813..3832)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3824..4639
                     /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             3824..4639
                     /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(3897..3916)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 atcacgcatc ttcccgacaa cgcagaccgt tccgtggcaa agcaaaagtt caaaatcacc
       61 aactggtcca cctacaacaa agctctcatc aaccgtggct ccctcacttt ctggctggat
      121 gatggggcga ttcaggcctg gtatgagtca gcaacacctt cttcacgagg cagacctcag
      181 cgctcaaaga tgcaggggta aaagctaacc gcatctttac cgacaaggca tccggcagtt
      241 caacagatcg ggaagggctg gatttgctga ggatgaaggt ggaggaaggt gatgtcattc
      301 tggtgaagaa gctcgaccgt cttggccgcg acaccgccga catgatccaa ctgataaaag
      361 agtttgatgc tcagggtgta gcggttcggt ttattgacga cgggatcagt accgacggtg
      421 atatggggca aatggtggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt
      481 ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc
      541 gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat
      601 gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg
      661 cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg
      721 atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga
      781 ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc
      841 ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc
      901 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
      961 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt tagctcactc
     1021 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     1081 gcggataaca atttcacaca ggaaacagct atgaccatga ttacggattc actggccgtc
     1141 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
     1201 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     1261 cagttgcgca gcctgaatgg cgaatggcgc tttgcctggt ttccggcacc agaagcggtg
     1321 ccggaaagct ggctggagtg cgatcttcct gaggccgata ctgtcgtcgt cccctcaaac
     1381 tggcagatgc acggttacga tgcgcccatc tacaccaacg tgacctatcc cattacggtc
     1441 aatccgccgt ttgttcccac ggagaatccg acgggttgtt actcgctcac atttaatgtt
     1501 gatgaaagct ggctacagga aggccagacg cgaattattt ttgatggcgt tggaattagc
     1561 ttatcgactg cacggtgcac caatgcttct ggcgtcaggc agccatcgga agctgtggtg
     1621 atctcgatcc cgcgaaatta atacgactca ctatagggag accacaacgg tttccctcta
     1681 gaaataattt tgtttaactt taagaaggag atatacatat gtgataagat tccgtatgcg
     1741 cacccttagc gagaggttta tcattaaggt caacctctgg atgttgtttc ggcatcctgc
     1801 attgaatctg agttactgtc tgttttcctt gttggaacgg agagcatcgc ctgatgctct
     1861 ccgagccaac caggaaaccc gttttttctg acgtaagggt gcgcaacttt ctaagaattc
     1921 gagctctaga taagtaagat ccggctgcta acaaagcccg aaaggaagct gagttggctg
     1981 ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg
     2041 gttttttgct gaaaggagga actatatccg gatagtcacg tagcgatagc ggagtgtata
     2101 attcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     2161 taatggtttc ttagccgatg cccttgagag ccttcaaccc tgcagcaatg gcaacaacgt
     2221 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     2281 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     2341 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     2401 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     2461 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     2521 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     2581 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     2641 tttcgttcca ctgagcgtca gaccccttaa taagatgatc ttcttgagat cgttttggtc
     2701 tgcgcgtaat ctcttgctct gaaaacgaaa aaaccgcctt gcagggcggt ttttcgaagg
     2761 ttctctgagc taccaactct ttgaaccgag gtaactggct tggaggagcg cagtcaccaa
     2821 aacttgtcct ttcagtttag ccttaaccgg cgcatgactt caagactaac tcctctaaat
     2881 caattaccag tggctgctgc cagtggtgct tttgcatgtc tttccgggtt ggactcaaga
     2941 cgatagttac cggataaggc gcagcggtcg gactgaacgg ggggttcgtg catacagtcc
     3001 agcttggagc gaactgccta cccggaactg agtgtcaggc gtggaatgag acaaacgcgg
     3061 ccataacagc ggaatgacac cggtaaaccg aaaggcagga acaggagagc gcacgaggga
     3121 gccgccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc accactgatt
     3181 tgagcgtcag atttcgtgat gcttgtcagg ggggcggagc ctatggaaaa acggctttgc
     3241 cgcggccctc tcacttccct gttaagtatc ttcctggcat cttccaggaa atctccgccc
     3301 cgttcgtaag ccatttccgc tcgccgcagt cgaacgaccg agcgtagcga gtcagtgagc
     3361 gaggaagcgg aatatatcct gtatcacata ttctgctgac gcaccggtgc agcctttttt
     3421 ctcctgccac atgaagcact tcactgacac cctcatcagt gccaacatag taagccagta
     3481 tacactccgc tagcgctgag gtctgcctcg tgaagaaggt gttgctgact cataccaggc
     3541 ctgaatcgcc ccatcatcca gccagaaagt gagggagcca cggttgatga gagctttgtt
     3601 gtaggtggac cagttggtga ttttgaactt ttgctttgcc acggaacggt ctgcgttgtc
     3661 gggaagatgc gtgatctgat ccttcaactc agcaaaagtt cgatttattc aacaaagcca
     3721 cgttgtgtct caaaatctct gatgttacat tgcacaagat aaaaatatat catcatgaac
     3781 aataaaactg tctgcttaca taaacagtaa tacaaggggt gttatgagcc atattcaacg
     3841 ggaaacgtct tgctcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta
     3901 taaatgggct cgcgataatg tcgggcaatc aggtgcgaca atctatcgat tgtatgggaa
     3961 gcccgatgcg ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac
     4021 agatgagatg gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca
     4081 ttttatccgt actcctgatg atgcatggtt actcaccact gcgatccccg ggaaaacagc
     4141 attccaggta ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt
     4201 gttcctgcgc cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt
     4261 atttcgtctc gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt
     4321 tgatgacgag cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataagctttt
     4381 gccattctca ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt
     4441 tgacgagggg aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata
     4501 ccaggatctt gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg
     4561 gctttttcaa aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat
     4621 gctcgatgag tttttctaat cagaattggt taattggttg taacactggc agagcattac
     4681 gctgacttga cgggacggcg gctttgttga ataaatcgaa cttttgctga gttgaaggat
     4741 cag
//
