LOCUS       sequence-489546-        9027 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pLAM12 vector backbone containing Eco1 recombitron with MspRecT and 
            a donor in the ncRNA to target MSMEG_5894 gene of Mycobacterium 
            smegmatis mc2 155.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9027)
  AUTHORS   Gonzalez-Delgado A, Bonillo-Lopez L, Johnson MS, Knodlseder N, Ko 
            CC, Lekbach Y, Oh JH, Selvakumar H, Wold MC, Yu Z, Aragon V, 
            Gralnick JA, Guell M, Hatfull GF, Keitz BK, Koskella B, Mutalik VK, 
            van Pijkeren JP, Shipman SL
  TITLE     Genome editing of phylogenetically distinct bacteria using portable 
            retron-mediated recombineering
  JOURNAL   bioRxiv 2025.06.16.660010
REFERENCE   2  (bases 1 to 9027)
  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: "bioRxiv 
            2025.06.16.660010"
COMMENT     Sequence Label: pRECMYC-Eco1
FEATURES             Location/Qualifiers
     source          1..9027
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      6947..6993
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(7143..7162)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             7154..7969
                     /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
                     KPHAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHNLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(7227..7246)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      8300..8888
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8789..8808
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaacgaccga gcgcaacgcg tgagcccacc agctccgtaa gttcgggcgc tgtgtggctc
       61 gtacccgcgc attcaggcgg cagggggtct aacgggtcta aggcggcgtg tacggccgcc
      121 acagcggctc tcagcggccc ggaaacgtcc tcgaaacgac gcatgtgttc ctcctggttg
      181 gtacaggtgg ttgggggtgc tcggctgtcg ctggtgttcc accaccaggg ctcgacggga
      241 gagcggggga gtgtgcagtt gtggggtggc ccctcagcga aatatctgac ttggagctcg
      301 tgtcggacca tacaccggtg attaatcgtg gtctactacc aagcgtgagc cacgtcgccg
      361 acgaatttga gcagctctgg ctgccgtact ggccgctggc aagcgacgat ctgctcgagg
      421 ggatctaccg ccaaagccgc gcgtcggccc taggccgccg gtacatcgag gcgaacccaa
      481 cagcgctggc aaacctgctg gtcgtggacg tagaccatcc agacgcagcg ctccgagcgc
      541 tcagcgcccg ggggtcccat ccgctgccca acgcgatcgt gggcaatcgc gccaacggcc
      601 acgcacacgc agtgtgggca ctcaacgccc ctgttccacg caccgaatac gcgcggcgta
      661 agccgctcgc atacatggcg gcgtgcgccg aaggccttcg gcgcgccgtc gatggcgacc
      721 gcagttactc aggcctcatg accaaaaacc ccggccacat cgcctgggaa acggaatggc
      781 tccactcaga tctctacaca ctcagccaca tcgaggccga gctcggcgcg aacatgccac
      841 cgccgcgctg gcgtcagcag accacgtaca aagcggctcc gacgccgcta gggcggaatt
      901 gcgcactgtt cgattccgtc aggttgtggg cctatcgtcc cgccctcatg cggatctacc
      961 tgccgacccg gaacgtggac ggactcggcc gcgcgatcta tgccgagtgc cacgcgcgaa
     1021 acgccgaatt tccgtgcaac gacgtgtgtc ccggaccgct accggacagc gaggtccgcg
     1081 ccatcgccaa cagcatttgg cgttggatca caaccaagtc gcgcatttgg gcggacggga
     1141 tcgtggtcta cgaggccaca ctcagtgcgc gccagtcggc catctcgcgg aagggcgcag
     1201 cagcgcgcac ggcggcgagc acagttgcgc ggcgcgcaaa gtccgcgtca gccatggagg
     1261 cattgctatg agcgacggct acagcgacgg ctacaaccgg cagccgactg tccgcaaaaa
     1321 gcggcgcgtg accgccgccg aaggcgctcg aatcaccgga ctatccgaac gccacgtcgt
     1381 ccggctcgtg gcgcaggaac gcagcgagtg gctcgccgag caggctgcac gccgcgaacg
     1441 catccgcgcc tatcacgacg acgagggcca ctcttggccg caaacggcca aacatttcgg
     1501 gctgcatctg gacaccgtta agcgactcgg ctatcgggcg aggaaagagc gtgcggcaga
     1561 acaggaagcg gctcaaaagg cccacaacga agccgacaat ccaccgctgt tctaacgcaa
     1621 ttggggagcg ggtgtcgcgg gggttccgtg gggggttccg ttgcaacggg tcggacaggt
     1681 aaaagtcctg gtagacgcta gttttctggt ttgggccatg cctgtctcgt tgcgtgtttc
     1741 gttgcgcccg ttttgaatac cagccagacg agacggggtt ctacgaatct tggtcgatac
     1801 caagccattt ccgctgaata tcggggagct caccgccaga atcggtggtt gtggtgatgt
     1861 acgtggcgaa ctccgttgta gtgcctgtgg tggcatccgt ggcgcggccg cggtaccaga
     1921 tctttaaatc tagataaaga agtgacgcgg tctcaagcgt cgagcgtcgc cagcgtgtcg
     1981 aggatgtcga agtcgtagcc gtcggcgctg gcgatgtaga cctgctggtc gaattgactg
     2041 tcgcgcatac acatcgggcc ccggggcccg tcgaacccga catcgtgcgc ggatgccatc
     2101 aggtccggta tctcggggga gtgggcccgc tggaagatgg cctcgagcgc aagcagaccc
     2161 tcgtaacagg attcggccat cgcgttgagc ggtggcgcgt cggcgccgta gcgggcgacg
     2221 tagctgccca tcaggtccat ggcacccgcg gtggccagtg aactgaagta cgccgcggcg
     2281 acatagaggt tttcggtgga gccggcgccg ctggccagca gcatgttctc ctccatcagc
     2341 gggctgaacc gcgccatgcg gtcgtgcccg ccggcgcgcg cgaactcgcg gttgaacaac
     2401 acggcgtcct ggccgacgag cagcatcaac acggcctgcg cccccgacgc gatggccttg
     2461 cggacaggtg cgcggaaatc gtcggtgccg tacgggacgt agatctcccg tctgagctcg
     2521 aggtccagat ctcggcagta cgcgcgggcg gccgcggcgg aacggcgcgg ccagatgtag
     2581 tcatcgccga ccaggcacca ggaccggatg ccgaagtggt cgcgcagcca ggcgagcgcg
     2641 ggcgcgatct ggatctgcgg tgtctcgcct gtgcagaaca cgcccggtgt gcgttcaccg
     2701 ccctcgtaca acgaggtgta gacgtacggg atgcggtcgc ggaccaccgg ggagatgcgg
     2761 ttgcgcacgg ccgagatgtg ccagccggtc acggcgtcga gaccgtgacc tcgcaaccgg
     2821 tcggcgacgg tccgggcgac gtcgtcgccg ggcgctccgc cgtcgagcac ctcgatggtg
     2881 accttgcggc cctgcaggcc gcctcggtcg ttgacctcct tggccgcgag ctcggccacg
     2941 gcctcgcacg aaggcgcgaa gattcccgct ggcccttgaa gcggaatcac cagcccgacg
     3001 cggaactcaa cctcgccgtc ctgcactcca gatcaccgtc gatcccgtgt agtctgcgct
     3061 tcaaagcttt ctagcagaaa taattcattc tgaacagacc ccgccgtcga cacgaggaga
     3121 cacccaccat ggccgccgga cagcagcgcc gccccaacct cctgctgccg ttggtgcgtc
     3181 tgacccacct cgcggagtcg gcgatcgaac gcgtgctcgc ggactcgtcg ctcaagatcg
     3241 aggactggcg cgtgctcgac gagttggccg gacggcgcac cgtgcccatg agcgatctcg
     3301 cgcaggccac gctgatcacg ggtccgactc tcaccagaac cgtcgatcgc cttgtgtcgc
     3361 aagggatcat ctaccggact gccgatctgc atgaccgccg gcgggtgctc gtggcgttga
     3421 ccccgcgggg gcggacgctg cgcaaccgcc tggtggacgc ggtagccgag gccgagtgtg
     3481 cggcttttga atcgtgcggg ctggacgtcg accagttgcg cgaactcgtc gacaccacct
     3541 cgaatttgac ttcgtaacca cccgcgcccg gcgcgggcgt tcacccttga cttttatttt
     3601 catctggata tatttcgggt gaatggaaag gggtgaccat gccgacctac acattccgtt
     3661 gttcccactg cggtcccttc gatctcacct gcgcgatctc cgagcgcgat gcggcggcga
     3721 cctgtccgga gtgccggacg ccggcgcgcc gggtcttcgg ttcggtaggg ctgacgacat
     3781 tcaccgcggg acatcaccgc gcattcgacg cggcgtccgc gagcgccgaa agtcccacgg
     3841 tggtgaagtc gattcccgca ggcgcggacc gcccgcgggc cccgcgccgc aatcccggtc
     3901 taccgagtct gccgaggtac tagcgacatg ggtggcgtcg ggctcttcta cgtgggtgcg
     3961 gtgctcatca tcgacgggct gatgctgctg ggccgcatca gcccacgagg cgcaacaccg
     4021 ctgaacttct tcgtcggcgg actgcaggtg gtgacgccta cggtgctgat cctgcagtcc
     4081 ggcggagacg cggccgtgat cttcgcggcc tccgggctct acctgttcgg cttcacctac
     4141 ctgtgggtgg ccatcaacaa cgtgaccgac tgggacggag aaggtctcgg atggttctcg
     4201 ctgttcgtcg cgatcgccgc actcggctac tcgtggcacg cgttcaccgc cgaggccgac
     4261 ccggcgttcg gggtgatctg gctgctgtgg gcagtgctgt ggttcatgct gttcctgctg
     4321 ctcggcctgg ggcacgacgc actggggccc gccgtcgggt tcgtcgcggt ggccgaaggc
     4381 gtgatcaccg ccgccgtgcc ggccttcctg atcgtgtcgg gcaactggga aaccggcccg
     4441 ctccccgccg cggtcatcgc cgtgatcggt tttgccgcag ttgttctcgc ataccccatc
     4501 gggcgccgtc tcgcagcgcc gtcagtcacc aaccctccac cggccgcgct cgcggccacc
     4561 acccgataag agaaagggag tccacatatg aagtcggccg agtacctgaa cacgttccgg
     4621 ctccgcaatc tcggcctgcc cgtgatgaat aacctgcacg acatgagcaa agccacccgc
     4681 atctcggtcg aaacgctgcg gctgctgatc tacacggccg attttcgcta tcgcatctat
     4741 accgtggaaa aaaagggccc agagaagcgt atgcgtacga tttaccagcc gtcgcgtgag
     4801 ttgaaggccc tccaaggttg ggtgctgcgt aatattttgg acaagctgag ctcgtccccc
     4861 ttctcgattg gttttgagaa gcaccaaagc attttgaaca acgccacgcc gcacatcggt
     4921 gccaatttca ttctcaatat cgacctcgaa gatttcttcc cctccctgac ggcgaataag
     4981 gtgtttggtg tctttcattc cttgggttac aaccggctga tctcgtccgt gctcacgaag
     5041 atttgttgct ataagaatct gctcccgcaa ggggccccgt ccagccccaa attggcgaac
     5101 ctcatctgta gcaaattgga ctaccgcatc cagggctatg cgggctcgcg cgggctgatc
     5161 tacacgcgct acgccgatga cctgacgctc tccgcgcaaa gcatgaaaaa ggtcgtgaaa
     5221 gcccgcgatt tcctctttag cattattccg agcgagggct tggtcattaa tagcaagaaa
     5281 acctgtatca gcggtccgcg cagccagcgg aaggtgacgg gcctggtgat ctcgcaagaa
     5341 aaggtgggta tcggccggga aaagtataaa gaaattcgtg ccaaaatcca tcacattttt
     5401 tgcgggaaat cgtcggagat tgaacacgtc cgggggtggt tgagctttat cctctcggtc
     5461 gattccaaaa gccatcggcg gctgattacg tacatcagca aactggagaa aaagtacggc
     5521 aaaaatccac tcaacaaggc gaagacgtaa catttaacta tgataagatt ccgtatgcgc
     5581 acccttagcg agaggtttat cattaaggtc aacctctgga tgttgtttcg gcatcctgca
     5641 ttgaatctga gttactgtct gttttcctga attctgcgcc gtcgtgcgtc acgcccggcg
     5701 gggcccgtgt gagccgctgg acaccatcac cccggcgcca gagggaattc aggaaacccg
     5761 ttttttctga cgtaagggtg cgcatacgga atctcggcgg gagaaaggag aaagccaatc
     5821 atggctgaaa atgctgtcac caagcaggat tcgcccaagg cacccgagac gatctcgcaa
     5881 gtgctgcagg tgctcgtgcc gcagctggcg cgtgcagtgc ccaagggtat ggaccccgac
     5941 cgcatcgcgc ggatcgtgca aaccgagatc cgcaagtcgc gcaacgcgaa agctgctggc
     6001 atcgctaagc agtccctcga cgactgcacg caagagtcat ttgccggtgc gctgctgacc
     6061 tcggccgcgc tcggtctcga gcccggtgtc aacggcgagt gctacctcgt gccctaccgc
     6121 gacacccggc gcggcgtggt cgagtgccag ctgatcatcg gctaccaggg catcgtcaaa
     6181 ctgttctggc agcacccgcg cgcctcccgg atcgacgcgc agtgggtcgg cgcgaacgac
     6241 gaattccatt acaccatggg cctcaatccg acgctgaaac acgtgaaggc caagggtgat
     6301 cggggtaatc cggtctactt ctacgcgatc gtcgaggtga ccggcgctga gccgctgtgg
     6361 gacgtgttca ccgccgacga gatcagggaa ttgcgtcgcg gcaaggtcgg atcctcgggc
     6421 gacatcaagg acccgcagcg ctggatggag cggaagaccg cgctcaaaca ggtgctgaag
     6481 ctggcaccga agacgacgcg gctcgacgcg gcgatccgcg ccgacgatcg cccgggcacc
     6541 gacctgtcac agtcgcaggc gctcgcgctg ccgtcgaccg tcaagccgac ggccgactac
     6601 atcgacggcg agatcgccga gccgcacgag gtcgatacgc caccgaagtc gtcgcgcgca
     6661 caacgcgcgc agcgcgccac cgcgccggcg cccgacgtgc agatggccaa tcccgatcag
     6721 ctgaagcgcc tcggcgagat ccagaaggcc gaaaagtaca acgacgccga ttggttcaag
     6781 ttcctcgccg attcggccgg cgtcaaagcc acgcgcgccg ccgacctcac gttcgacgag
     6841 gcgaaggctg tcatcgacat gttcgacggg cccaacgcat gagcgccccg gcgaattccg
     6901 acgcggtggt tgatctgcaa ctagcgtacg atcgactgcc aggcatcaaa taaaacgaaa
     6961 ggctcagtcg aaagactggg cctttcgttt tatgccatca tggccgcggt gatcagctag
     7021 ccacctgacg tcgggggggg gggaaagcca cgttgtgtct caaaatctct gatgttacat
     7081 tgcacaagat aaaaatatat catcatgaac aataaaactg tctgcttaca taaacagtaa
     7141 tacaaggggt gttatgagcc atattcaacg ggaaacgtct tgctcgaggc cgcgattaaa
     7201 ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc
     7261 aggtgcgaca atctatcgct tgtatgggaa gccccatgcg ccagagttgt ttctgaaaca
     7321 tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac
     7381 ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt
     7441 actcaccact gcgatccccg ggaaaacagc attccaggta ttagaagaat atcctgattc
     7501 aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt
     7561 ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat
     7621 gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga
     7681 acaagtctgg aaagaaatgc ataatctttt gccattctca ccggattcag tcgtcactca
     7741 tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga
     7801 tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct
     7861 cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc
     7921 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaat cagaattggt
     7981 taattggttg taacactggc agagcattac gctgacttga cgggacggcg gctttgttga
     8041 ataaatcgaa cttttgctga gttgaaggat cagatcacgc atcttcccga caacgcagac
     8101 cgttccgtgg caaagcaaaa gttcaaaatc accaactggt ccacctacaa caaagctctc
     8161 accaaccgtg gctccctcac tttctggctg gatgatgggg cgattcaggc ctggtatgag
     8221 tcagcaacac cttcttcacg aggcagacct cactagttcc atgagcgtca gaccccgtag
     8281 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     8341 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     8401 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     8461 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     8521 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     8581 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     8641 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     8701 gcgccacgct tcccgagggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     8761 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     8821 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     8881 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     8941 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     9001 agtgagctga taccgctcgc cgcagcc
//
