LOCUS       sequence-444676-        3130 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Destination vector with kanamycin resistance and pBR322 origin of 
            replication. Carries G-H CIDAR MoClo overhangs for level 1 construct
            cloning..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3130)
  AUTHORS   Nielsen JR, Lewis MJ, Huang WE
  TITLE     Construction and Characterization of MoClo-Compatible Vectors for 
            Modular Protein Expression in E. coli.
  JOURNAL   ACS Synth Biol. 2025 Jan 12. doi: 10.1021/acssynbio.4c00564.
  PUBMED    39801078
REFERENCE   2  (bases 1 to 3130)
  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: "ACS Synth 
            Biol. 2025 Jan 12. doi: 10.1021/acssynbio.4c00564."
COMMENT     Sequence Label: DVK_pBR322_GH
FEATURES             Location/Qualifiers
     source          1..3130
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     327..346
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     411..430
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     624..642
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(646..689)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    692..713
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     CDS             complement(708..1031)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCRSLAMSS
                     TSL"
     primer_bind     926..948
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     940..957
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     941..957
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    958..1014
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(1027..1043)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1027..1043)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1040..1062)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1051..1067
                     /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(join(1075..1081,1082..1099,1100..1105))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1120..1141
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             1452..1643
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQKKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1637..1659)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    1745..1884
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     1800..1819
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1899..1916)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2070..2658)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2150..2169)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2689..2728)
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     CDS             complement(join(2734..3130,1..419))
                     /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             complement(join(2734..3130,1..419))
                     /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"
ORIGIN
        1 ggacaattac aaacaggaat cgaatgcaac cggcgcagga acactgccag cgcatcaaca
       61 atattttcac ctgaatcagg atattcttct aatacctgga atgctgtttt cccggggatc
      121 gcagtggtga gtaaccatgc atcatcagga gtacggataa aatgcttgat ggtcggaaga
      181 ggcataaatt ccgtcagcca gtttagtctg accatctcat ctgtaacatc attggcaacg
      241 ctacctttgc catgtttcag aaacaactct ggcgcatcgg gcttcccata caatcgatag
      301 attgtcgcac ctgattgccc gacattatcg cgagcccatt tatacccata taaatcagca
      361 tccatgttgg aatttaatcg cggcctggag caagacgttt cccgttgaat atggctcata
      421 acaccccttg tattactgtt tatgtaagca gacagtttta ttgttcatga tgatatattt
      481 ttatcttgtg caatgtaaca tcagagattt tgagacacaa cgtggctttg ttgaataaat
      541 cgaacttttg ctgagttgaa ggatcagctc gagtgccacc tgacgtctaa gaaaccatta
      601 ttatcatgac attaacctat aaaaataggc gtatcacgag gcagaatttc agataaaaaa
      661 aatccttagc tttcgctaag gatgatttct ggaattcgcg gccgcttcta gagactagtg
      721 gaagacattg ccagagacct gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg
      781 agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
      841 tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga
      901 ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgaa
      961 ttcgagctcg gtacccgggg atcctctaga gtcgacctgc aggcatgcaa gcttggcgta
     1021 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     1081 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     1141 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     1201 atgaatcggc caacgcgcgg gggtctctac taatgtcttc taacacctac atctgtatta
     1261 acgaagcgct ggcattgacc ctgagtgatt tttctctggt cccgccgcat ccataccgcc
     1321 agttgtttac cctcacaacg ttccagtaac cgggcatgtt catcatcagt aacccgtatc
     1381 gtgagcatcc tctctcgttt catcggtatc attaccccat gaacagaaat cccccttaca
     1441 cggaggcatc agtgaccaaa cagaaaaaaa ccgcccttaa catggcccgc tttatcagaa
     1501 gccagacatt aacgcttctg gagaaactca acgagctgga cgcggatgaa caggcagaca
     1561 tctgtgaatc gcttcacgac cacgctgatg agctttaccg cagctgcctc gcgcgtttcg
     1621 gtgatgacgg tgaaaacctc tgacacatgc agctcccggc gacggtcaca gcttgtctgt
     1681 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     1741 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc
     1801 ggcatcagag cagattgtac tgagagtgca ccattgcggt gtgaaatacc gcacagatgc
     1861 gtaaggagaa aataccgcat caggcgctct tccgcttcct cgctcactga ctcgctgcgc
     1921 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     1981 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     2041 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     2101 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     2161 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     2221 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     2281 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     2341 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     2401 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     2461 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt
     2521 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     2581 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     2641 agaaaaaaag gatctcaaga agatcctttg atcttttcta ccggtccgtc caaaaaaaaa
     2701 ggctccaaaa ggagccttta attgtatcgg tgattagaaa aactcatcga gcatcaaatg
     2761 aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa gccgtttctg
     2821 taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct ggtatcggtc
     2881 tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt caaaaataag
     2941 gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg gcaaaagctt
     3001 atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat caaaatcact
     3061 cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcaagacgaa atacgcgatc
     3121 gctgttaaaa
//
