LOCUS       sequence-435738-        7487 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 0 pAHP6-PhiC31-NLS-tUBQ1 construct for assembly of two 
            integrase constructs, GG0 and GG4 BsaI sites..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7487)
  TITLE     Nemhauser Lab Plasmids 2024
REFERENCE   2  (bases 1 to 7487)
  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: L0I97 - GG0-pAHP6-PhiC31-NLS-tUBQ1-GG4
FEATURES             Location/Qualifiers
     source          1..7487
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(39..833)
                     /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             complement(39..833)
                     /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(150..169)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     760..779
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(1182..1484)
                     /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     complement(1284..1303)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     1607..1629
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1621..1638
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1622..1638
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1645..1664
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1645..1663
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3344..5158
                     /codon_start=1
                     /product="integrase from phage phi-C31"
                     /label=phage phi-C31 integrase
                     /translation="MDTYAGAYDRQSRERENSSAASPATQRSANEDKAADLQREVERDG
                     GRFRFVGHFSEAPGTSAFGTAERPEFERILNECRAGRLNMIIVYDVSRFSRLKVMDAIP
                     IVSELLALGVTIVSTQEGVFRQGNVMDLIHLIMRLDASHKESSLKSAKILDTKNLQREL
                     GGYVGGKAPYGFELVSETKEITRNGRMVNVVINKLAHSTTPLTGPFEFEPDVIRWWWRE
                     IKTHKHLPFKPGSQAAIHPGSITGLCKRMDADAVPTRGETIGKKTASSAWDPATVMRIL
                     RDPRIAGFAAEVIYKKKPDGTPTTKIEGYRIQRDPITLRPVELDCGPIIEPAEWYELQA
                     WLDGRGRGKGLSRGQAILSAMDKLYCECGAVMTSKRGEESIKDSYRCRRRKVVDPSAPG
                     QHEGTCNVSMAALDKFVAERIFNKIRHAEGDEETLALLWEAARRFGKLTEAPEKSGERA
                     NLVAERADALNALEELYEDRAAGAYDGPVGRKHFRKQQAALTLRQQGAEERLAELEAAE
                     APKLPLDQWFPEDADADPTGPKSWWGRASVDDKRVFVGLFVDKIVVTKSTTGRGQGTPI
                     EKRASITWAKPPTDDDEDDAQDGTEDVAA"
     CDS             5159..5179
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5818..5834)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5818..5834)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5831..5853)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5842..5858
                     /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(5866..5872,5873..5890,5891..5896))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5911..5932
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6049..6066)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6220..6808)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6300..6319)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6946..7320)
                     /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"
     regulatory      7317..7326
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 agaaaatacc gcatcaggaa attgtaagcg ttaataattc agaagaactc gtcaagaagg
       61 cgatagaagg cgatgcgctg cgaatcggga gcggcgatac cgtaaagcac gaggaagcgg
      121 tcagcccatt cgccgccaag ctcttcagca atatcacggg tagccaacgc tatgtcctga
      181 tagcggtccg ccacacccag ccggccacag tcgatgaatc cagaaaagcg gccattttcc
      241 accatgatat tcggcaagca ggcatcgcca tgggtcacga cgagatcctc gccgtcgggc
      301 atgctcgcct tgagcctggc gaacagttcg gctggcgcga gcccctgatg ctcttcgtcc
      361 agatcatcct gatcgacaag accggcttcc atccgagtac gtgctcgctc gatgcgatgt
      421 ttcgcttggt ggtcgaatgg gcaggtagcc ggatcaagcg tatgcagccg ccgcattgca
      481 tcagccatga tggatacttt ctcggcagga gcaaggtgag atgacaggag atcctgcccc
      541 ggcacttcgc ccaatagcag ccagtccctt cccgcttcag tgacaacgtc gagcacagct
      601 gcgcaaggaa cgcccgtcgt ggccagccac gatagccgcg ctgcctcgtc ttgcagttca
      661 ttcagggcac cggacaggtc ggtcttgaca aaaagaaccg ggcgcccctg cgctgacagc
      721 cggaacacgg cggcatcaga gcagccgatt gtctgttgtg cccagtcata gccgaatagc
      781 ctctccaccc aagcggccgg agaacctgcg tgcaatccat cttgttcaat catgcgaaac
      841 gatcctcatc ctgtctcttg atcagagctt gatcccctgc gccatcagat ccttggcggc
      901 aagaaagcca tccagtttac tttgcagggc ttcccaacct taccagaggg cgccccagct
      961 ggcaattccg gttcgcttgc tgtccataaa accgcccagt ctagctatcg ccatgtaagc
     1021 ccactgcaag ctacctgctt tctctttgcg cttgcgtttt cccttgtcca gatagcccag
     1081 tagctgacat tcatccgggg tcagcaccgt ttctgcggac tggctttcta cgtgaaaagg
     1141 atctaggtga agatcctttt tgataatctc atgcctgaca tttatattcc ccagaacatc
     1201 aggttaatgg cgtttttgat gtcattttcg cggtggctga gatcagccac ttcttccccg
     1261 ataacggaga ccggcacact ggccatatcg gtggtcatca tgcgccagct ttcatccccg
     1321 atatgcacca ccgggtaaag ttcacgggag actttatctg acagcagacg tgcactggcc
     1381 agggggatca ccatccgtcg ccccggcgtg tcaataatat cactctgtac atccacaaac
     1441 agacgataac ggctctctct tttataggtg taaaccttaa actgccgtac gtataggctg
     1501 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa
     1561 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt
     1621 tgtaaaacga cggccagtga attgtaatac gactcactat agggcgaatt gggccctcta
     1681 gatgcatgct cgagcggccg ccagtgtgat ggatatctgc agaattcagg cactctgtgg
     1741 tctcaggagc acggggcgca aagaagcatg acatacgaat gagacaattt cagttttcgt
     1801 atataataat tccataatca taatttaatc actagtttta attggtaaca taaccttaaa
     1861 aaagcatcca ctcattatgc gctccacttt actacttact ctgataacat gaaaaagtat
     1921 atgtttaata ctagctagtt gtttgttccg tttccaagca cgctctttct tcttttcagt
     1981 catttttaat ccgcatttac aataatattt agctattttc atatatcatt ctatactagc
     2041 tcacaataat tacatatgag cacgcttttt tgtgttcaaa tattataaat cttcgttagt
     2101 taccgctgct ttttataatt atgtatactt tctggttcca ggagactaat taaggaagat
     2161 tattagaggc ttcttatacg tgccatatga ttatttattt atgtactgtc atggttcata
     2221 tatagcgatg tgataccaat ataccaatca ccaaagttat gagaaaatca cgtaaccaga
     2281 ggtgttggta catacatttt ctgttaaaaa tagttaagag gacttgtaat cttttgtgtt
     2341 taactattga taatttttta ctttattcgg tgaaacaatt tacgcatgtt tctgtgttgt
     2401 gcttgtgttt cttgtattct tcttgtgtat agctttcctt ataagaaaga aaaagaagaa
     2461 gattaattga ttagatcgag aaagagaaag gccttttgag atagaaatgg agaaaggatg
     2521 gattgaattt gaagcataga gaatagcgaa cactcactgc aaagttagtg ggaaagtcgc
     2581 gcgtcatacg agggagcaac aaagtgtggg cttttctttt acagtctcag aaactaatgc
     2641 atgctcacat ttaaaaaaaa aaaaaaaaaa acaattcttg tgttaaatca agagtggatt
     2701 aattatgaga taatctaatg tacattactg ttatatgatt ataacttgac agaccaaata
     2761 atcatcttaa tcagacctga taaaagaaaa gagattaatt tcaacagtga gaaattaaag
     2821 atagaaagta catattattg gtgtaaaagt tgttgattta atttaaggca gatgtattaa
     2881 gttaaatatg taaatgtaga ggctgcggga agggtgacca gctggtctga cagggtacgc
     2941 cggttgtcgg gaggaagaac gccgctattg gcagagtcac gcggggtaac cggctggcgg
     3001 ctgtagccgg tcacagagtg tattaatatg gtcgtacaat acttgtagag tgctttttcc
     3061 tgtgacggtc ccgtttattc cccacaccct cctcgattct ttcacttaaa agtgggggaa
     3121 aaaggtaacc tgactcttag aaaatttgaa tacaattgtg ggtcacctca tctcatgtag
     3181 tcatgtctat acaattataa atacctctca acaaacccaa aatcgaatta taagccttct
     3241 ctagctagaa tttgaagcaa gacctgttca cattagtaaa aacaaatata tagaaagagg
     3301 aaagattgaa gcggaaaacg ggacaagacg ggtgtgccgt tgaatggaca cgtatgccgg
     3361 cgcttatgat agacaatcaa gagaaagaga gaactcttct gccgcctctc cagctacaca
     3421 gagatctgcc aacgaggata aagctgctga tttacaaaga gaggtagaga gagatggggg
     3481 taggtttaga tttgtgggtc atttctctga ggctcctggt acatcagcct ttggtactgc
     3541 tgaacggcct gaatttgaaa gaatattaaa tgagtgtcgg gccggtcgac taaatatgat
     3601 aatagtttat gatgtatcta ggttcagtag actcaaagtt atggatgcaa ttccgatcgt
     3661 gagcgagcta cttgcactcg gtgttactat cgtaagtaca caagaaggtg tattcaggca
     3721 gggtaatgtg atggacctta tacaccttat tatgcgactc gacgctagcc ataaagagtc
     3781 atcactgaag agtgctaaaa tcttggacac caagaacctg caaagggagc ttggaggcta
     3841 tgtaggagga aaagctccgt atggtttcga actcgtgagt gagacaaaag agattaccag
     3901 aaatggtagg atggtgaacg tggttattaa taagctagct catagcacaa cccctctcac
     3961 agggcctttt gagttcgaac ctgacgttat tagatggtgg tggagagaga tcaaaactca
     4021 caaacatctt ccatttaagc caggtagcca ggctgctatc caccctggat ctatcactgg
     4081 gctttgtaaa aggatggatg ctgacgcagt gccaactcga ggggaaacaa tcggtaagaa
     4141 gactgcatct tccgcttggg atcccgctac agttatgaga atccttcggg atccgcgtat
     4201 tgctggattc gccgcagaag tcatttataa gaaaaaacct gatggtactc ctactacaaa
     4261 aatcgaaggt tataggatcc aaagggatcc tattactttg cgtccagttg aattagactg
     4321 tggaccgatc atcgaacctg ccgagtggta cgagttgcag gcctggcttg atggtagagg
     4381 gcgtggaaag ggtttatctc ggggacaggc aattttgtca gcaatggata agctttattg
     4441 tgagtgtggg gctgttatga catcaaaacg tggagaagag tcaatcaagg attcttacag
     4501 gtgcagacgt cgtaaagtgg tggacccttc tgctccagga cagcatgagg gtacatgcaa
     4561 cgtttctatg gctgctcttg ataaatttgt ggccgaacgt attttcaaca agattagaca
     4621 cgctgaaggg gatgaggaaa ctttagctct tctttgggag gctgcccggc gttttggtaa
     4681 gctaacagag gcaccagaaa aaagtggcga aagagctaat ttggtagccg aaagagcaga
     4741 cgccttgaat gcattggaag aactttacga agatagggct gccggagcat atgatggacc
     4801 agttggtcgg aagcatttca ggaagcaaca agctgcacta actttgagac agcaaggcgc
     4861 agaggagcga ctcgcagagc ttgaggccgc cgaggcccca aagcttccac tcgatcaatg
     4921 gtttcctgag gatgccgacg ctgatcctac aggtccaaag agctggtggg gaagagctag
     4981 cgtagacgat aagagagtgt ttgtgggtct atttgttgac aagattgtag ttactaagtc
     5041 cactacgggt cgaggacagg gcactcccat cgaaaagaga gccagtatta cctgggcaaa
     5101 gcctccaacc gatgatgatg aagacgatgc acaagatggt accgaggatg tggcagcacc
     5161 aaaaaaaaag aggaaggtgt aagcttagag actcttatca agaatcccat ctcttgcttg
     5221 cttttttttg ttgtcttccc tttgataggg tttgtttttc ttgtttcagt gactttctat
     5281 gttaaaagat aatgtcagta aaaggatttg gttttctatt attctgaatc gattacggaa
     5341 gattcttgct taattccaat ctatacaagt atcgtgaaat aatgaccgtt tatgtgatta
     5401 ggagacgtgt ttcattaata aaatataaga tcaatacatt gttagtagtg ataaactatg
     5461 tacaaattgt attgattgta aaagaaacac aataggttcc ttttttctac aatatattgt
     5521 gacagactct ctgttttaac gaatgaatta aatttgtcaa cgaaaaaatt aaagaatgat
     5581 gtgatcactt tgcaccttgg ttcatcttca gaggcttctg catgttccat gcattgaggc
     5641 tctctacggt cacaggttgt gagccgttgt taaagacaaa caaatggctt tgagacctca
     5701 tcacgcctga attccagcac actggcggcc gttactagtg gatccgagct cggtaccaag
     5761 cttgatgcat agcttgagta ttctaacgcg tcacctaaat agcttggcgt aatcatggtc
     5821 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     5881 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     5941 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     6001 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     6061 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     6121 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     6181 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     6241 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     6301 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     6361 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     6421 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     6481 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     6541 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     6601 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     6661 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     6721 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     6781 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     6841 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     6901 cttcacctag atccttttaa attaaaaatg aagttttagc acgtgtcagt cctgctcctc
     6961 ggccacgaag tgcacgcagt tgccggccgg gtcgcgcagg gcgaactccc gcccccacgg
     7021 ctgctcgccg atctcggtca tggccggccc ggaggcgtcc cggaagttcg tggacacgac
     7081 ctccgaccac tcggcgtaca gctcgtccag gccgcgcacc cacacccagg ccagggtgtt
     7141 gtccggcacc acctggtcct ggaccgcgct gatgaacagg gtcacgtcgt cccggaccac
     7201 accggcgaag tcgtcctcca cgaagtcccg ggagaacccg agccggtcgg tccagaactc
     7261 gaccgctccg gcgacgtcgc gcgcggtgag caccggaacg gcactggtca acttggccat
     7321 ggtggccctc ctcacgtgct attattgaag catttatcag ggttattgtc tcatgagcgg
     7381 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     7441 aaaagtgcca cctgatgcgg tgtgaaatac cgcacagatg cgtaagg
//
