LOCUS       sequence-435737-        5252 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 0 pMUTE promoter, GG0 and GG1 BsaI sites for golden gate 
            assembly..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5252)
  TITLE     Nemhauser Lab Plasmids 2024
REFERENCE   2  (bases 1 to 5252)
  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"
COMMENT     Sequence Label: L0I89-pMUTE_GG0-1
FEATURES             Location/Qualifiers
     source          1..5252
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1807..1826)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1808..1826)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1833..1850)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1833..1849)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1842..1864)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1987..2289
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2168..2187
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             2638..3432
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2638..3432
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2692..2711)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3302..3321
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      3632..3641
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3638..4012
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     rep_origin      4150..4738
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4639..4658
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4892..4909
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5026..5047
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5062..5067,5068..5085,5086..5092)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5100..5116
                     /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     5105..5127
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5124..5140
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5124..5140
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 cactctgtgg tctcaggagc caaaccaatc ccatacccaa accagactga aaattggttg
       61 gtttggtttt cgacccttgc attgaatgtt aaccgatatc ttgatcgaac gtgagttttt
      121 tttttttttt ttttcttgtt tcattgcctt gtcgatcaat attgtatata acgtgataga
      181 ccatgaacaa attgaatatc gaagtaggag ttagatttct ttactcgtaa ttaaatgggg
      241 catgcacctt gcatgtctta tacgtattgt agcctaacat ttgcggctac aaacatttgt
      301 caccaacatt ttcattccag ctgcttccac agttttggga cttcggagat aatgtatatg
      361 tggatatgca tatatgaacc ataaaaataa acgtttgtcc aaacaaaacc acaaatagaa
      421 gtgttatcca agtgggtttg tttcatagaa aaaatattgt cactatagca atttttttgg
      481 ttaggttaaa ccttatttaa tgaagtttgt aatatgaatt gagatattgt taactttgca
      541 aaacagagcc gaaattttac gaatatctct ttattaaaaa taaataatat aatttaagaa
      601 tacttctcgg gaggaatctt aatgagccgt ccaggtagct gaacaactga actttatctt
      661 tgctccaaag gttatatcaa tttatgtttt cttgtagaac ctatatttca tgcaaaaagt
      721 tatccattaa ccaatggaaa ttggttccga ctagtgatca aggaaaatga tggaatcatc
      781 acgaatgaga aatttgtggt attaaatgac atagtgatta aggaaaacaa aaagttgtaa
      841 ttcaactaca aactatgtcg taactattta tattttgtta cttgtcaatg ttgtcattta
      901 tcttacaaag tcaataggat agaagatgtg aaacataatt acacaaaaac gatgtcatat
      961 agaaacatgc gaaatcacat aatcatttca tcacttcgaa atttcgtaga tttccaatca
     1021 gatactaccc aatatgtttg ctaatagtaa gtagtaaaaa acatatgtta cgccaaaaaa
     1081 aacggctttt gatgtactta ttttccttca tcgatccatg aaaactgtaa aacatctcaa
     1141 tacaaactaa acctttccac cagtatatgt ccgattcaat gtttttttac tttgtgtatg
     1201 tatcagtgta ttatctgtta ataaagagta aaagaaatcc attttttctt gtgaggacac
     1261 gcggtaatta agaacaaaga agtggcacaa gtgtagaaat taaactagaa gtaaaagaag
     1321 gtggtgcaaa ggcatgcact ctaaatgaaa atacgatatt gaaattcata aaagacctca
     1381 cgcagtgaat gtaaatgtaa atgtatccaa cttcacaaac gtgacgtatc tttgtgtatg
     1441 catacccttt tctttacgtt gttctaatac atatcgatag taataagaga gagaaagaga
     1501 aaaatcaaga agagacaaaa gaatgtcacg agacagcgac atagtgcatt tatgaacctt
     1561 gaaaagaaat tctaaaccct aattatcact tgttgttgcg gctgtataaa tacgactttg
     1621 ttttgaagaa tgtgtcgaac aaagtgaaaa cataagatca tcttcttcgt tgatagatca
     1681 atataggaac tccagaagag aatcttgatc aattaagtat caatgtgaga ccatcaacgt
     1741 cctgaattct gcagatatcc atcacactgg cggccgctcg agcatgcatc tagagggccc
     1801 aattcgccct atagtgagtc gtattacaat tcactggccg tcgttttaca acgtcgtgac
     1861 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
     1921 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctatac
     1981 gtacggcagt ttaaggttta cacctataaa agagagagcc gttatcgtct gtttgtggat
     2041 gtacagagtg atattattga cacgccgggg cgacggatgg tgatccccct ggccagtgca
     2101 cgtctgctgt cagataaagt ctcccgtgaa ctttacccgg tggtgcatat cggggatgaa
     2161 agctggcgca tgatgaccac cgatatggcc agtgtgccgg tctccgttat cggggaagaa
     2221 gtggctgatc tcagccaccg cgaaaatgac atcaaaaacg ccattaacct gatgttctgg
     2281 ggaatataaa tgtcaggcat gagattatca aaaaggatct tcacctagat ccttttcacg
     2341 tagaaagcca gtccgcagaa acggtgctga ccccggatga atgtcagcta ctgggctatc
     2401 tggacaaggg aaaacgcaag cgcaaagaga aagcaggtag cttgcagtgg gcttacatgg
     2461 cgatagctag actgggcggt tttatggaca gcaagcgaac cggaattgcc agctggggcg
     2521 ccctctggta aggttgggaa gccctgcaaa gtaaactgga tggctttctt gccgccaagg
     2581 atctgatggc gcaggggatc aagctctgat caagagacag gatgaggatc gtttcgcatg
     2641 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     2701 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     2761 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa
     2821 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     2881 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     2941 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     3001 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     3061 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     3121 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc
     3181 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     3241 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     3301 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     3361 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     3421 gagttcttct gaattattaa cgcttacaat ttcctgatgc ggtattttct ccttacgcat
     3481 ctgtgcggta tttcacaccg catcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     3541 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     3601 ataaatgctt caataatagc acgtgaggag ggccaccatg gccaagttga ccagtgccgt
     3661 tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag ttctggaccg accggctcgg
     3721 gttctcccgg gacttcgtgg aggacgactt cgccggtgtg gtccgggacg acgtgaccct
     3781 gttcatcagc gcggtccagg accaggtggt gccggacaac accctggcct gggtgtgggt
     3841 gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc gtgtccacga acttccggga
     3901 cgcctccggg ccggccatga ccgagatcgg cgagcagccg tgggggcggg agttcgccct
     3961 gcgcgacccg gccggcaact gcgtgcactt cgtggccgag gagcaggact gacacgtgct
     4021 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     4081 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     4141 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     4201 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     4261 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     4321 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     4381 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     4441 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     4501 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     4561 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     4621 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     4681 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     4741 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     4801 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     4861 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
     4921 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
     4981 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
     5041 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
     5101 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagctatt
     5161 taggtgacgc gttagaatac tcaagctatg catcaagctt ggtaccgagc tcggatccac
     5221 tagtaacggc cgccagtgtg ctggaattca gg
//
