LOCUS       sequence-435864-        4178 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 0 tUBQ10 terminator, GG4 and GG6 BsaI sites for golden gate 
            assembly..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4178)
  TITLE     Nemhauser Lab Plasmids 2024
REFERENCE   2  (bases 1 to 4178)
  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: L0I74-_tUBQ10_GG4-GG6
FEATURES             Location/Qualifiers
     source          1..4178
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(733..752)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(734..752)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(759..776)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(759..775)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(768..790)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             913..1215
                     /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     1094..1113
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1564..2358
                     /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             1564..2358
                     /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(1618..1637)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2228..2247
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2558..2567
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2564..2938
                     /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      3076..3664
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3565..3584
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3818..3835
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3952..3973
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3988..3993,3994..4011,4012..4018)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4026..4042
                     /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     4031..4053
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4050..4066
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4050..4066
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 cactctgtgg tctcagctta tctcgtctct gttatgctta agaagttcaa tgtttcgttt
       61 catgtaaaac tttggtggtt tgtgttttgg ggccttgtat aatccctgat gaataagtgt
      121 tctactatgt ttccgttcct gttatctctt tctttctaat gacaagtcga acttcttctt
      181 tatcatcgct tcgtttttat tatctgtgct tcttttgttt aatacgcctg caaagtgact
      241 cgactctgtt tagtgcagtt ctgcgaaact tgtaaatagt ccaattgttg gcctctagta
      301 atagatgtag cgaaagtgtt gagctgttgg gttctaagga tggcttgaac atgttaatct
      361 tttaggttct gagtatgatg aacattcgtt gttgctaaga aatgcctgta atgtcccaca
      421 aatgtagaaa atggttcgta cctttgtcca agcattgata tgtctgatga gaggaaactg
      481 caagatactg agcttggttt aacgaaggag aggcagtttc ttccttccaa agcatttcat
      541 ttgacaatgc cttgatcatc ttaagtagag tttctgttgt ggaaagtttg aaactttgaa
      601 gaaacgactc tcaagtaaat tgatgatcac aagtgaaagt gtatgttcgc ttgagacctc
      661 cgatcccctg aattctgcag atatccatca cactggcggc cgctcgagca tgcatctaga
      721 gggcccaatt cgccctatag tgagtcgtat tacaattcac tggccgtcgt tttacaacgt
      781 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
      841 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
      901 ctatacgtac ggcagtttaa ggtttacacc tataaaagag agagccgtta tcgtctgttt
      961 gtggatgtac agagtgatat tattgacacg ccggggcgac ggatggtgat ccccctggcc
     1021 agtgcacgtc tgctgtcaga taaagtctcc cgtgaacttt acccggtggt gcatatcggg
     1081 gatgaaagct ggcgcatgat gaccaccgat atggccagtg tgccggtctc cgttatcggg
     1141 gaagaagtgg ctgatctcag ccaccgcgaa aatgacatca aaaacgccat taacctgatg
     1201 ttctggggaa tataaatgtc aggcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     1261 ttcacgtaga aagccagtcc gcagaaacgg tgctgacccc ggatgaatgt cagctactgg
     1321 gctatctgga caagggaaaa cgcaagcgca aagagaaagc aggtagcttg cagtgggctt
     1381 acatggcgat agctagactg ggcggtttta tggacagcaa gcgaaccgga attgccagct
     1441 ggggcgccct ctggtaaggt tgggaagccc tgcaaagtaa actggatggc tttcttgccg
     1501 ccaaggatct gatggcgcag gggatcaagc tctgatcaag agacaggatg aggatcgttt
     1561 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     1621 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     1681 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     1741 ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     1801 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     1861 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     1921 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     1981 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     2041 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc
     2101 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     2161 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     2221 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     2281 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     2341 cttgacgagt tcttctgaat tattaacgct tacaatttcc tgatgcggta ttttctcctt
     2401 acgcatctgt gcggtatttc acaccgcatc aggtggcact tttcggggaa atgtgcgcgg
     2461 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     2521 accctgataa atgcttcaat aatagcacgt gaggagggcc accatggcca agttgaccag
     2581 tgccgttccg gtgctcaccg cgcgcgacgt cgccggagcg gtcgagttct ggaccgaccg
     2641 gctcgggttc tcccgggact tcgtggagga cgacttcgcc ggtgtggtcc gggacgacgt
     2701 gaccctgttc atcagcgcgg tccaggacca ggtggtgccg gacaacaccc tggcctgggt
     2761 gtgggtgcgc ggcctggacg agctgtacgc cgagtggtcg gaggtcgtgt ccacgaactt
     2821 ccgggacgcc tccgggccgg ccatgaccga gatcggcgag cagccgtggg ggcgggagtt
     2881 cgccctgcgc gacccggccg gcaactgcgt gcacttcgtg gccgaggagc aggactgaca
     2941 cgtgctaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     3001 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     3061 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     3121 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     3181 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     3241 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     3301 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     3361 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     3421 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     3481 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     3541 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     3601 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     3661 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     3721 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     3781 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     3841 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
     3901 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
     3961 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
     4021 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa
     4081 gctatttagg tgacgcgtta gaatactcaa gctatgcatc aagcttggta ccgagctcgg
     4141 atccactagt aacggccgcc agtgtgctgg aattcagg
//
