LOCUS       sequence-344705-        5572 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of J106 scRNA on pBBR1-KmR plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5572)
  AUTHORS   Kiattisewee C, Dong C, Fontana J, Sugianto W, Peralta-Yahya P, 
            Carothers JM, Zalatan JG
  TITLE     Portable bacterial CRISPR transcriptional activation enables 
            metabolic engineering in Pseudomonas putida.
  JOURNAL   Metab Eng. 2021 Apr 27. pii: S1096-7176(21)00063-X. doi: 
            10.1016/j.ymben.2021.04.002.
  PUBMED    33930546
REFERENCE   2  (bases 1 to 5572)
  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: "Metab Eng. 
            2021 Apr 27. pii: S1096-7176(21)00063-X. doi: 
            10.1016/j.ymben.2021.04.002."
COMMENT     Sequence Label: pPPC009.106
FEATURES             Location/Qualifiers
     source          1..5572
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1822..2616
                     /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(1876..1895)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2486..2505
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    3021..3042
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3057..3062,3063..3080,3081..3087)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3095..3111
                     /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     3100..3122
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3119..3135
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3119..3135
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3154..3174
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3156..3174
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     terminator      complement(3219..3265)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3418..3435
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     CDS             complement(3491..3508)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(3524..3553)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     misc_RNA        complement(3576..3594)
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     promoter        complement(3678..3712)
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     primer_bind     complement(3726..3745)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3727..3745)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3755..3772)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3755..3771)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3764..3786)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(4480..5142)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      join(5143..5572,1..342)
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
ORIGIN
        1 cttcgactgc ccccactcgc ataggcttgg gtcgttccag gcgcgtcaag gccaagccgc
       61 tgcgcggtcg ctgcgcgagc cttgacccgc cttccacttg gtgtccaacc ggcaagcgaa
      121 gcgcgcaggc cgcaggccgg aggcttttcc ccagagaaaa ttaaaaaaat tgatggggca
      181 aggccgcagg ccgcgcagtt ggagccggtg ggtatgtggt cgaaggctgg gtagccggtg
      241 ggcaatccct gtggtcaagc tcgtgggcag gcgcagcctg tccatcagct tgtccagcag
      301 ggttgtccac gggccgagcg aagcgagcca gccggtggcc gctcgcggcc atcgtccaca
      361 tatccacggg ctggcaaggg agcgcagcga ccgcgcaggg cgaagcccgg agagcaagcc
      421 cgtagggcgc cgcagccgcc gtaggcggtc acgactttgc gaagcaaagt ctagtgagta
      481 tactcaagca ttgagtggcc cgccggaggc accgccttgc gctgcccccg tcgagccggt
      541 tggacaccaa aagggagggg caggcatggc ggcatacgcg atcatgcgat gcaagaagct
      601 ggcgaaaatg ggcaacgtgg cggccagtct caagcacgcc taccgcgagc gcgagacgcc
      661 caacgctgac gccagcagga cgccagagaa cgagcactgg gcggccagca gcaccgatga
      721 agcgatgggc cgactgcgcg agttgctgcc agagaagcgg cgcaaggacg ctgtgttggc
      781 ggtcgagtac gtcatgacgg ccagcccgga atggtggaag tcggccagcc aagaacagca
      841 ggcggcgttc ttcgagaagg cgcacaagtg gctggcggac aagtacgggg cggatcgcat
      901 cgtgacggcc agcatccacc gtgacgaaac cagcccgcac atgaccgcgt tcgtggtgcc
      961 gctgacgcag gacggcaggc tgtcggccaa ggagttcatc ggcaacaaag cgcagatgac
     1021 ccgcgaccag accacgtttg cggccgctgt ggccgatcta gggctgcaac ggggcatcga
     1081 gggcagcaag gcacgtcaca cgcgcattca ggcgttctac gaggccctgg agcggccacc
     1141 agtgggccac gtcaccatca gcccgcaagc ggtcgagcca cgcgcctatg caccgcaggg
     1201 attggccgaa aagctgggaa tctcaaagcg cgttgagacg ccggaagccg tggccgaccg
     1261 gctgacaaaa gcggttcggc aggggtatga gcctgcccta caggccgccg caggagcgcg
     1321 tgagatgcgc aagaaggccg atcaagccca agagacggcc cgagaccttc gggagcgcct
     1381 gaagcccgtt ctggacgccc tggggccgtt gaatcgggat atgcaggcca aggccgccgc
     1441 gatcatcaag gccgtgggcg aaaagctgct gacggaacag cgggaagtcc agcgccagaa
     1501 acaggcccag cgccagcagg aacgcgggcg cgcacatttc cccgaaaagt gccacctggg
     1561 atgaatgtca gctactgggc tatctggaca agggaaaacg caagcgcaaa gagaaagcag
     1621 gtagcttgca gtgggcttac atggcgatag ctagactggg cggttttatg gacagcaagc
     1681 gaaccggaat tgccagctgg ggcgccctct ggtaaggttg ggaagccctg caaagtaaac
     1741 tggatggctt tcttgccgcc aaggatctga tggcgcaggg gatcaagatc tgatcaagag
     1801 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     1861 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     1921 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     1981 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     2041 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     2101 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     2161 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     2221 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     2281 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     2341 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     2401 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     2461 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     2521 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     2581 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga
     2641 ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg
     2701 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     2761 atctcatgct ggagttcttc gcccaccccc atgggcaaat attatacgca aggcgacaag
     2821 gtgctgatgc cgctggcgat tcaggttcat catgccgttt gtgatggctt ccatgtcggc
     2881 agaatgctta atgaattaca acagttttta tgcatgcgcc caatacgcaa accgcctctc
     2941 cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg
     3001 ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta
     3061 cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca
     3121 ggaaacagct atgaccatga ttacgccaag cgcgcaatta accctcacta aagggaacaa
     3181 aagctggtct agcttaagag ttcaccgaca aacaacagat aaaacgaaag gcccagtctt
     3241 tcgactgagc ctttcgtttt atttgatgcc tggcagttcc ctactctcgc atggggagac
     3301 cccacactac catcggcgct acggcgtttc acttctgagt tcggcatggg gtcaggtggg
     3361 accaccgcgc tactgccgcc aggcaaattc tgttttatca gaccgcttct gcgttctgat
     3421 ttaatctgta tcaggctgaa aatcttctct catccgccaa aacagccaag ctggagaccg
     3481 tttaaactca atgatgatga tgatgatggt cgacggcgct attcagatcc tcttctgaga
     3541 tgagtttttg ttcgggccca agcttcaaaa aaagcacatg ggtgatcctc atgtgccact
     3601 ttttcaagtt gataacggac tagccttatt ttaacttgct atttctagct ctaaaacgcg
     3661 gttaccaaag gcgtcctact agtattatac ctaggactga gctagctgtc aagagctcca
     3721 attcgcccta tagtgagtcg tattacgcgc gctcactggc cgtcgtttta caacgtcgtg
     3781 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca
     3841 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga
     3901 atggcgaatg gaaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     3961 aatcagctca ttttttaacc aataggccgt actgcgatga gtggcagggc ggggcgtaat
     4021 ttttttaagg cagttattgg tgcccttaaa cgcctggttg ctacgcctga ataagtgata
     4081 ataagcggat gaatggcaga aattcgaaag caaattcgac ccggtcgtcg gttcagggca
     4141 gggtcgttaa atagccgctt atgtctattg ctggtttacc ggtttattga ctaccggaag
     4201 cagtgtgacc gtgtgcttct caaatgcctg aggccagttt gctcaggctc tccccgtgga
     4261 ggtaataatt gacgatatga tcatttattc tgcctcccag agcctgataa aaacggtgaa
     4321 tccgttagcg aggtgccgcc ggcttccatt caggtcgagg tggcccggct ccatgcaccg
     4381 cgacgcaacg cggggaggca gacaaggtat agggcggcga ggcggctaca gccgatagtc
     4441 tggaacagcg cacttacggg ttgctgcgca acccaagtgc taccggcgcg gcagcgtgac
     4501 ccgtgtcggc ggctccaacg gctcgccatc gtccagaaaa cacggctcat cgggcatcgg
     4561 caggcgctgc tgcccgcgcc gttcccattc ctccgtttcg gtcaaggctg gcaggtctgg
     4621 ttccatgccc ggaatgccgg gctggctggg cggctcctcg ccggggccgg tcggtagttg
     4681 ctgctcgccc ggatacaggg tcgggatgcg gcgcaggtcg ccatgcccca acagcgattc
     4741 gtcctggtcg tcgtgatcaa ccaccacggc ggcactgaac accgacaggc gcaactggtc
     4801 gcggggctgg ccccacgcca cgcggtcatt gaccacgtag gccgacacgg tgccggggcc
     4861 gttgagcttc acgacggaga tccagcgctc ggccaccaag tccttgactg cgtattggac
     4921 cgtccgcaaa gaacgtccga tgagcttgga aagtgtcttc tggctgacca ccacggcgtt
     4981 ctggtggccc atctgcgcca cgaggtgatg cagcagcatt gccgccgtgg gtttcctcgc
     5041 aataagcccg gcccacgcct catgcgcttt gcgttccgtt tgcacccagt gaccgggctt
     5101 gttcttggct tgaatgccga tttctctgga ctgcgtggcc atgcttatct ccatgcggta
     5161 ggggtgccgc acggttgcgg caccatgcgc aatcagctgc aacttttcgg cagcgcgaca
     5221 acaattatgc gttgcgtaaa agtggcagtc aattacagat tttctttaac ctacgcaatg
     5281 agctattgcg gggggtgccg caatgagctg ttgcgtaccc ccctttttta agttgttgat
     5341 ttttaagtct ttcgcatttc gccctatatc tagttctttg gtgcccaaag aagggcaccc
     5401 ctgcggggtt cccccacgcc ttcggcgcgg ctccccctcc ggcaaaaagt ggcccctccg
     5461 gggcttgttg atcgactgcg cggccttcgg ccttgcccaa ggtggcgctg cccccttgga
     5521 acccccgcac tcgccgccgt gaggctcggg gggcaggcgg gcgggcttcg cc
//
