LOCUS       sequence-418202-        3543 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Type TU2-b backbone with kanamycin resistance and ColE1-bom-rop ORI.
            amilCP dropout vector..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3543)
  AUTHORS   Shapiro M
  TITLE     EcoFlex MoClo Extension, Volume I
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 3543)
  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: 
            "unpublished"
COMMENT     Sequence Label: pTU2-b_CrK_amilCP
FEATURES             Location/Qualifiers
     source          1..3543
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(126..920)
                     /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
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEDHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(237..256)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     847..866
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     terminator      complement(1059..1102)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     promoter        1125..1159
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     RBS             1168..1179
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      1852..1923
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      complement(1946..2040)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             2354..2545
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2539..2561)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2645..2784
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2700..2719
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2792..2809)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(2963..3543,1..8))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3043..3062)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtga acgaaaactc
       61 acgttaaagc aagcaaagaa aaaaggcgac agattactct gtcgcctttt ttcctgactc
      121 ataactcaga agaactcgtc aagaaggcga tagaaggcga tgcgctgcga atcgggagcg
      181 gcgataccgt aaagcacgag gaagcggtca gcccattcgc cgccaagttc ttcagcaata
      241 tcacgggtag ccaacgctat gtcctgatag cggtccgcca cacccagccg gccacagtcg
      301 atgaatccag aaaagcggcc attttccacc atgatattcg gcaagcaggc atcgccgtgg
      361 gtcacgacga gatcctcgcc gtcgggcata cgcgccttga gcctggcgaa cagttcggct
      421 ggcgcgagcc cctgatgatc ttcgtccaga tcatcctgat cgacaagacc ggcttccatc
      481 cgagtacgtg ctcgctcgat gcgatgtttc gcttggtggt cgaatgggca ggtagccgga
      541 tcaagcgtat gcagccgccg cattgcatca gccatgatgg atactttctc ggcaggagca
      601 aggtgagatg acaggagatc ctgccccggc acttcgccca atagcagcca gtcccttccc
      661 gcttcagtga caacgtcgag cacagccgcg caaggaacgc ccgtcgtggc cagccacgat
      721 agccgcgctg cctcgtcttg cagttcattc agggcaccgg acaggtcggt cttgacaaaa
      781 agaaccgggc gaccctgcgc tgacagccgg aacacggcgg catcagagca gccgattgtc
      841 tgttgtgccc agtcatagcc gaatagcctc tccacccaag ccgccggaga acctgcgtgc
      901 aatccatctt gttcaatcat gcgaaacgat cctcatcctg tctcttgatc agatcttgat
      961 cccctgcgcc atcagatcct tggcggcaag aaagccatcc agtttacttt gcagggcttc
     1021 ccaaccttac cagagggcgc cccagctggc aattccggat ttcagataaa aaaaatcctt
     1081 agctttcgct aaggatgatt tcggtctcac tatatctaga gacgtttaca gctagctcag
     1141 tcctaggtat tatgctagct actagagaaa gaggagaaat actagatgag tgtgatcgct
     1201 aaacaaatga cctacaaggt ttatatgtca ggcacggtca atggacacta ctttgaggtc
     1261 gaaggcgatg gaaaaggtaa gccctacgag ggggagcaga cggtaaagct cactgtcacc
     1321 aagggcggac ctctgccatt tgcttgggat attttatcac cacagtgtca gtacggaagc
     1381 ataccattca ccaagtaccc tgaagatatc cctgactatg taaagcagtc attcccggag
     1441 ggctatacat gggagaggat catgaacttt gaagatggtg cagtgtgtac tgtcagcaat
     1501 gattccagca tccaaggcaa ctgtttcatc taccatgtca agttctctgg tttgaacttt
     1561 cctcccaatg gacctgtcat gcagaagaaa acacagggct gggaacccaa cactgagcgt
     1621 ctgtttgcac gagatggaat gctgctagga aacaacttta tggctctgaa gttagaagga
     1681 ggcggtcact atttgtgtga atttaaaact acttacaagg caaagaagcc tgtgaagatg
     1741 ccagggtatc actatgttga ccgcaaactg gatgtaacca atcacaacaa ggattacact
     1801 tcggttgagc agtgtgaaat ttccattgca cgcaaacctg tggtcgccta acaaataaaa
     1861 cgaaaggctc agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct
     1921 ctccgtctca gaaggtacag agaccattct caccaataaa aaacgcccgg cggcaaccga
     1981 gcgttctgaa caaatccaga tggagttctg aggtcattac tggatctatc aacaggagtc
     2041 ctgcggtttc cgtgtttcgt aaagtctgga aacgcggaag tcagcgccct gcaccattat
     2101 gttccggatc tgcatcgcag gatgctgctg gctaccctgt ggaacaccta catctgtatt
     2161 aacgaagcgc tggcattgac cctgagtgat ttttctctgg tcccgccgca tccataccgc
     2221 cagttgttta ccctcacaac gttccagtaa ccgggcatgt tcatcatcag taacccgtat
     2281 cgtgagcatc ctctctcgtt tcatcggtat cattaccccc atgaacagaa atccccctta
     2341 cacggaggca tcagtgacca aacaggaaaa aaccgccctt aacatggccc gctttatcag
     2401 aagccagaca ttaacgcttc tggagaaact caacgagctg gacgcggatg aacaggcaga
     2461 catctgtgaa tcgcttcacg accacgctga tgagctttac cgcagctgcc tcgcgcgttt
     2521 cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gacggtcaca gcttgtctgt
     2581 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     2641 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc
     2701 ggcatcagag cagattgtac tgagagtgca ccattgcggt gtgaaatacc gcacagatgc
     2761 gtaaggagaa aataccgcat caggccgctt cctcgctcac tgactcgctg cgctcggtcg
     2821 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     2881 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2941 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     3001 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     3061 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     3121 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     3181 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     3241 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     3301 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     3361 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct
     3421 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     3481 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     3541 aag
//
