LOCUS       40924_46713        5960 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pWW3351, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5960)
  AUTHORS   Whitaker WR, Shepherd ES, Sonnenburg JL.
  TITLE     Tunable expression tools enable single-cell strain distinction in 
            the gut microbiome
  JOURNAL   Cell (2017) In press
REFERENCE   2  (bases 1 to 5960)
  AUTHORS   Whitaker WR, Shepherd ES, Sonnenburg JL.
  TITLE     Direct Submission
  JOURNAL   
REFERENCE   3  (bases 1 to 5960)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5960)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell (2017)
            In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (15-MAR-2017) Microbiology "
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5960
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(5..393)
                     /direction=LEFT
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K;
                     requires the R6K initiator protein pi for replication"
     oriT            564..673
                     /label=oriT
                     /note="incP origin of transfer"
     regulatory      817..909
                     /regulatory_class="promoter"
     regulatory      914..998
                     /regulatory_class="ribosome_binding_site"
     CDS             996..1709
                     /label=superfolder GFP
                     /note="GFP variant that folds robustly even when fused to
                     poorly folded proteins (Pedelacq et al., 2006)"
     CDS             1716..1739
                     /label=FLAG
                     /note="FLAG(R) epitope tag, followed by an enterokinase
                     cleavage site"
     CDS             1740..1763
                     /label=FLAG
                     /note="FLAG(R) epitope tag, followed by an enterokinase
                     cleavage site"
     CDS             1764..1787
                     /label=FLAG
                     /note="FLAG(R) epitope tag, followed by an enterokinase
                     cleavage site"
     CDS             1791..1808
                     /label=6xHis
                     /note="6xHis affinity tag"
     terminator      1831..1858
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     CDS             complement(2090..3373)
                     /codon_start=1
                     /product="NBU integrase"
                     /label=NBU integrase
                     /protein_id="ARI71329.1"
                     /translation="MNIKRNIIFALESRKKNGVPIVENVPIRMRVIFASQRIEFTTGYR
                     IDVAKWDADKQRVKSGCTNKLKQSAAEINTDLLKYYAEIQNIFKEFEVQEVMPTTQQLK
                     EAFNMRMKDTSEEQPEEAPVSFWEVFDEFVKECGNQNNWTASTYEKFAAVRNHLKEFKE
                     DATFNYFNEFGLNEYVNFLRDTKDMRNSTIGKQMGFLKWFLRWSFKKGHHQNIAYDTFK
                     PKLKTTSKKVIFLTWDELNKLKDYQIPKDKQYLERVRDVFLFCCFTSLRYSDVRNLKRS
                     DVKSDHIEITTVKTADSLTIELNKYSKAILDKYKDIHFENYMALPVISNQKMNDYLKEL
                     GELAEINEPVRETYYKGNERIDEVTPKYALLSTHAGRRTFICNALALGIPAQVVMKWTG
                     HSDYKAMKPYIDIADDIKANAMNKFNQL"
     misc_feature    3533..3544
                     /label=integration site
                     /note="integration site"
     CDS             complement(3816..4673)
                     /label=AmpR
                     /note="beta-lactamase"
     CDS             5161..5895
                     /codon_start=1
                     /product="ErmG"
                     /label=ErmG
                     /protein_id="ARI71332.1"
                     /translation="MNKVNIKDSQNFITSKYHIEKIMNCISLDEKDNIFEIGAGKGHFT
                     AGLVKRCNFVTAIEIDSKLCEVTRNKLLNYPNYQIVNDDILKFTFPSHNPYKIFGSIPY
                     NISTNIIRKIVFESSATISYLIVEYGFAKMLLDTNRSLALLLMAEVDISILAKIPRYYF
                     HPKPKVDSTLIVLKRKPAKMAFKERKKYETFVMKWVNKEYEKLFTKNQFNKALKHARIY
                     DINNISFEQFVSLFNSYKIFNG"
ORIGIN
        1 cccatgtcag ccgttaagtg ttcctgtgtc actcaaaatt gctttgagag gctctaaggg
       61 cttctcagtg cgttacatcc ctggcttgtt gtccacaacc gttaaacctt aaaagcttta
      121 aaagccttat atattctttt ttttcttata aaacttaaaa ccttagaggc tatttaagtt
      181 gctgatttat attaatttta ttgttcaaac atgagagctt agtacgtgaa acatgagagc
      241 ttagtacgtt agccatgaga gcttagtacg ttagccatga gggtttagtt cgttaaacat
      301 gagagcttag tacgttaaac atgagagctt agtacgtgaa acatgagagc ttagtacgta
      361 ctatcaacag gttgaactgc tgatcttcag atcctctacg ccggacgcat cgtggccgga
      421 tcaattccgt tttccgctgc ataaccctgc ttcggggtca ttatagcgat tttttcggta
      481 tatccatcct ttttcgcacg atatacagga ttttgccaaa gggttcgtgt agactttcct
      541 tggtgtatcc aacggcgtca gccgggcagg ataggtgaag taggcccacc cgcgagcggg
      601 tgttccttct tcactgtccc ttattcgcac ctggcggtgc tcaacgggaa tcctgctctg
      661 cgaggctggc cggctaccgc cggcgtaaca gatgagggca agcggatggc tgatgaaacc
      721 aagccaacca ggaagggcag cccacctatc acggaattga tccccctcga attggaattc
      781 gttccgtctc gattcagatc taaaggatcc aatctagagt ttgcaatggt taatctattg
      841 ttaaaattta aagtttcact tgaactttca aataatgttc ttatatttgc agtgtcgaaa
      901 gaaacaaagg actgatcaca caacatttaa aaaataacat tatgaaagca ctcgaaaaga
      961 gattcaaagg taatattaac aataatttat tttcaatgcg taaaggcgaa gagctgttca
     1021 ctggtgtcgt ccctattctg gtggaactgg atggtgatgt caacggtcat aagttttccg
     1081 tgcgtggcga gggtgaaggt gacgcaacta atggtaaact gacgctgaag ttcatctgta
     1141 ctactggtaa actgccggtt ccttggccga ctctggtaac gacgctgact tatggtgttc
     1201 agtgctttgc tcgttatccg gaccatatga agcagcatga cttcttcaag tccgccatgc
     1261 cggaaggcta tgtgcaggaa cgcacgattt cctttaagga tgacggcacg tacaaaacgc
     1321 gtgcggaagt gaaatttgaa ggcgataccc tggtaaaccg cattgagctg aaaggcattg
     1381 actttaaaga agacggcaat atcctgggcc ataagctgga atacaatttt aacagccaca
     1441 atgtttacat caccgccgat aaacaaaaaa atggcattaa agcgaatttt aaaattcgcc
     1501 acaacgtgga ggatggcagc gtgcagctgg ctgatcacta ccagcaaaac actccaatcg
     1561 gtgatggtcc tgttctgctg ccagacaatc actatctgag cacgcaaagc gttctgtcta
     1621 aagatccgaa cgagaaacgc gatcatatgg ttctgctgga gttcgtaacc gcagcgggca
     1681 tcacgcatgg tatggatgaa ctgtacaaag gatctgatta caaggacgat gatgataagg
     1741 actacaaaga tgatgatgac aaagattata aggacgatga tgataaaggt catcatcatc
     1801 atcatcacta aatcctagct gattagctcg agaaggccat cctgacggat ggcctttttt
     1861 ttgtatcgag acgactagtg taattgccta tcttccagtg atggaacagc atttgtgcat
     1921 tggctgcaac aatcagcctt acttgtgcct gttctatttc cgaaccgacc gcttgtatga
     1981 atccatcaaa attcgttttc tctatgttgg attccttgtt gctcatattg tgatgataat
     2041 ttctacaaat atagtcattg gtaactatct atgaaactgt ttgatacttt tatagttgat
     2101 taaacttgtt catggcattt gccttaatat catccgctat gtcaatgtag ggtttcatag
     2161 ctttgtagtc gctgtgtccc gtccatttca tgaccacctg tgccgggatt ccgagagcca
     2221 gcgcattgca gatgaatgtc cttcttcctg catgggtact gagcaaagcg tatttgggtg
     2281 tgacttcatc aatacgttca tttcccttgt agtaggtttc ccgtacaggc tcgttgattt
     2341 ctgccagttc gcccagctct ttcaggtaat cgttcatctt ctggttgctg atgacgggca
     2401 gagccatgta attctcgaaa tggatgtcct tgtatttgtc cagtatggct ttgctgtatt
     2461 tgttcagttc aatcgtcagg ctgtcggcag tcttgactgt ggttatttcg atgtggtcgg
     2521 acttcacatc gcttcttttc agattgcgaa catccgaata ccgcaaactc gtaaagcagc
     2581 agaacaggaa aacatcacgc acacgttcca ggtattgctt atccttgggt atctggtagt
     2641 ctttcagctt gttcagttca tcccaagtca ggaagattac ttttttcgag gtggttttca
     2701 gtttcggttt gaacgtatcg tatgcaatgt tctgatgatg tcctttcttg aagctccagc
     2761 gcaggaacca tttgaggaat cccatttgct tgccgatggt gctgtttctc atatccttgg
     2821 tgtcacgcag gaagttgacg tattcgttca atccaaactc gttgaaatag ttgaacgttg
     2881 catcctcctt gaactctttg aggtggttcc tcactgctgc aaatttttca taggtggatg
     2941 ccgtccagtt attctggtta ccgcactctt ttacaaactc atcgaacacc tcccaaaagc
     3001 tgacaggggc ttcttccggc tgttcttcgc tggtgtcttt cattctcatg ttgaaagctt
     3061 ccttcaactg ttgggtcgtt ggcatgacct cctgcacctc aaattccttg aaaatattct
     3121 ggatttcggc atagtatttc agcaagtccg tattgatttc ggctgcactt tgctttagct
     3181 tgttggtaca tccgctcttt acccgctgct tatctgcatc ccatttggct acgtcaatcc
     3241 ggtagcccgt tgtaaactcg atgcgttggc tggcaaagat gacacgcata cggatgggta
     3301 cgttctctac gattggcaca ccgttctttt tccggctctc caatgcaaaa atgatgttgc
     3361 gcttgatatt cataattggg tgcgtttgaa attctacacc caaatataca cccaattatt
     3421 gagatagcaa aagacattta gaaacattta cttttactct atattgtaat ttacacttga
     3481 ttatcagtcg tttgcagtct tatgatattc tgtgaaagta taagttcgag agcctgtctc
     3541 tccgcaaaaa acgctgaaaa tcagcagatt gcaaaacaaa caccctgttt tacacccaag
     3601 aatgtaaagt cggctgtttt tgttttattt aagataatac aaccactaca taataaaaga
     3661 gtagcgatat taaaagaatc cgatgagaaa agactaatat ttatctatcc attcagtttg
     3721 atttttcagg actttacatc gtcctgaaag tatttgttgg taccgaggac gcgacagctg
     3781 cagacgcgtt atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     3841 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     3901 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgggaccca
     3961 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4021 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4081 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     4141 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     4201 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     4261 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     4321 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     4381 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     4441 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     4501 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     4561 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     4621 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     4681 ctttttcaat catggccgcg ggattaaaag tcggggattg gtgaacaaaa aggtgtttct
     4741 ctctttaaga gaaatatcgt tttgctaaac agttgatatt gaggtatcat tttatcgtaa
     4801 aagacatttt tgctcaacaa ttgcttgacg gaaatcaaca aattttagca ttttgtaaaa
     4861 aagtcgctat ataatttggt gaattggagt tattttcata tttttgcatc ccgaagagtt
     4921 tctcttaaag agagaaacat cttttgcata ccttttccga ccgaattttt atgtcgtaaa
     4981 gaggggcttt gcagggggtg gactcagaaa gatgagaata gatgactatt gtagttgaaa
     5041 cacatagaaa gttgctgata tacagaccga tacgcatatc gggatgaacc atgagtacgt
     5101 tcttttctca aaaaacataa atattcgaaa agagatgcaa taaattaagg agaggttata
     5161 atgaacaaag taaatataaa agatagtcaa aattttatta cttcaaaata tcacatagaa
     5221 aaaataatga attgcataag tttagatgaa aaagataaca tctttgaaat aggtgcaggg
     5281 aaaggtcatt ttactgctgg attggtaaag agatgtaatt ttgtaacggc gatagaaatt
     5341 gattctaaat tatgtgaggt aactcgtaat aagctcttaa attatcctaa ctatcaaata
     5401 gtaaatgatg atatactgaa atttacattt cctagccaca atccatataa aatatttggc
     5461 agcatacctt acaacataag cacaaatata attcgaaaaa ttgtttttga aagttcagcc
     5521 acaataagtt atttaatagt ggaatatggt tttgctaaaa tgttattaga tacaaacaga
     5581 tcactagcat tgctgttaat ggcagaggta gatatttcta tattagcaaa aattcctagg
     5641 tattatttcc atccaaaacc taaagtggat agcacattaa ttgtattaaa aagaaagcca
     5701 gcaaaaatgg catttaaaga gagaaaaaaa tatgaaactt ttgtaatgaa atgggttaac
     5761 aaagagtacg aaaaactgtt tacaaaaaat caatttaata aagctttaaa acatgcgaga
     5821 atatatgata taaacaatat tagtttcgaa caatttgtat cgctatttaa tagttataaa
     5881 atatttaacg gctaaaaaca ataggccaca tgcaactgta aatgtttacg cgggtaccga
     5941 caccgcggtg gaggggaatt
//
