LOCUS       SEGA078                6528 bp ds-DNA     linear   UNA 17-JUN-2021
DEFINITION  LMBP 12914.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 6528)
  AUTHORS   Morten Norholm Group
  TITLE     Direct Submission
  JOURNAL   Exported Jun 17, 2021 from SnapGene 4.2.11
COMMENT     SEGA landing pad: galK-em7-rrnB-PRha-low-BCD-GFP-tetA-S2P11-S3P22-rrnC
FEATURES             Location/Qualifiers
     source          1..6528
                     /organism="unspecified"
                     /mol_type="genomic DNA"
     source          605..1753
                     /organism="Escherichia coli str. K-12 substr. MG1655"
                     /strain="K-12"
                     /sub_strain="MG1655"
                     /mol_type="genomic DNA"
                     /db_xref="taxon:511145"
     CDS             1..528
                     /codon_start=3
                     /label=glmS
                     /translation="VHGLQALPSRIEQMLSQDKRIEALAEDFSDKHHALFLGRGDQYPI
                     ALEGALKLKEISYIHAEAYAAGELKHGPLALIDADMPVIVVAPNNELLEKLKSNIEEVR
                     ARGGQLYVFADQDAGFVSSDNMHIIEMPHVEEVIAPIFYTVPLQLLAYHVALIKGTDVD
                     QPRNLAKSVTVE"
     misc_feature    538..594
                     /label=L3S2P11 Terminator
     gene            complement(605..1753)
                     /gene="galK"
                     /gene_synonym="ECK0746; galA; JW0740"
                     /locus_tag="b0757"
                     /label=galK
                     /db_xref="EcoGene:EG10363"
                     /db_xref="GeneID:945358"
     CDS             complement(605..1753)
                     /codon_start=1
                     /transl_table=11
                     /gene="galK"
                     /gene_synonym="ECK0746; galA; JW0740"
                     /locus_tag="b0757"
                     /product="galactokinase"
                     /function="enzyme; Degradation of small molecules: Carbon 
                     compounds"
                     /EC_number="2.7.1.6"
                     /label=galK
                     /db_xref="GI:16128725"
                     /db_xref="ASAP:ABE-0002568"
                     /db_xref="UniProtKB/Swiss-Prot:P0A6T3"
                     /db_xref="EcoGene:EG10363"
                     /db_xref="GeneID:945358"
                     /protein_id="NP_415278.1"
                     /translation="MSLKEKTQSLFANAFGYPATHTIQAPGRVNLIGEHTDYNDGFVLP
                     CAIDYQTVISCAPRDDRKVRVMAADYENQLDEFSLDAPIVAHENYQWANYVRGVVKHLQ
                     LRNNSFGGVDMVISGNVPQGAGLSSSASLEVAVGTVLQQLYHLPLDGAQIALNGQEAEN
                     QFVGCNCGIMDQLISALGKKDHALLIDCRSLGTKAVSMPKGVAVVIINSNFKRTLVGSE
                     YNTRREQCETGARFFQQPALRDVTIEEFNAVAHELDPIVAKRVRHILTENARTVEAASA
                     LEQGDLKRMGELMAESHASMRDDFEITVPQIDTLVEIVKAVIGDKGGVRMTGGGFGGCI
                     VALIPEELVPAVQQAVAEQYEAKTGIKETLYVCKPSQGAGQC"
     gene            complement(605..611)
                     /gene="galM"
                     /gene_synonym="ECK0745; JW0739"
                     /locus_tag="b0756"
                     /label=galM
                     /db_xref="EcoGene:EG11698"
                     /db_xref="GeneID:944943"
     misc_feature    644..645
                     /label=F370L
     misc_feature    1754..1842
                     /label=Insert galK construct
     promoter        complement(1783..1830)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     terminator      1843..1886
                     /label=rrnB
                     /label=Terminator rrnB T1
     CDS             complement(1899..2744)
                     /codon_start=1
                     /gene="E. coli rhaR"
                     /product="transcriptional activator of rhaR and rhaS"
                     /label=rhaR
                     /note="needed for rhamnose-dependent activation of the rhaB
                     promoter"
                     /translation="MAHQLKLLKDDFFASDQQAVAVADRYPQDVFAEHTHDFCELVIVW
                     RGNGLHVLNDRPYRITRGDLFYIHADDKHSYASVNDLVLQNIIYCPERLKLNLDWQGAI
                     PGFNASAGQPHWRLGSMGMAQARQVIGQLEHESSQHVPFANEMAELLFGQLVMLLNRHR
                     YTSDSLPPTSSETLLDKLITRLAASLKSPFALDKFCDEASCSERVLRQQFRQQTGMTIN
                     QYLRQVRVCHAQYLLQHSRLLISDISTECGFEDSNYFSVVFTRETGMTPSQWRHLNSQK
                     D"
     CDS             complement(2818..3654)
                     /codon_start=1
                     /gene="E. coli rhaS"
                     /product="positive regulator of the rhaB promoter"
                     /label=rhaS
                     /note="activated by L-rhamnose"
                     /translation="MTVLHSVDFFPSGNASVAIEPRLPQADFPEHHHDFHEIVIVEHGT
                     GIHVFNGQPYTITGGTVCFVRDHDRHLYEHTDNLCLTNVLYRSPDRFQFLAGLNQLLPQ
                     ELDGQYPSHWRVNHSVLQQVRQLVAQMEQQEGENDLPSTASREILFMQLLLLLRKSSLQ
                     ENLENSASRLNLLLAWLEDHFADEVNWDAVADQFSLSLRTLHRQLKQQTGLTPQRYLNR
                     LRLMKARHLLRHSEASVTDIAYRCGFSDSNHFSTLFRREFNWSPRDIRQGRDGFLQ"
     promoter        3800..3918
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring 
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
     RBS             3930..3941
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     misc_feature    3942..3956
                     /label=TIR
     CDS             3948..4003
                     /codon_start=1
                     /label=BCD
                     /translation="MKAIFVLKHLNHAKEVF"
     misc_feature    3986..3994
                     /label=SD2
     misc_feature    3999..4001
                     /label=STOP BCD
     misc_feature    4001..4003
                     /label=start GOI
     CDS             4004..4714
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006) ?"
                     /label=GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKAN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     misc_feature    4718..4766
                     /label=L3S3P22
     promoter        4777..4805
                     /label=Promoter 3BCD2
     CDS             4845..6035
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMSGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSITGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     misc_feature    6016..6065
                     /label=HR3
     terminator      6046..6086
                     /label=rrnC
     misc_feature    6087..6136
                     /label=homology
     CDS             6281..6528
                     /codon_start=1
                     /label=pstS
                     /translation="MKVMRTTVATVVAATLSMSAFSVFAEASLTGAGATFPAPVYAKWA
                     DTYQKETGNKVNYQGIGSSGGVKQIIANTVDFGASDA"
ORIGIN
        1 gtgcatggtc tgcaggcgct gccgagccgt attgagcaga tgctgtctca ggacaaacgc
       61 attgaagcgc tggcagaaga tttctctgac aaacatcacg cgctgttcct gggccgtggc
      121 gatcagtacc caatcgcgct ggaaggcgca ttgaagttga aagagatctc ttacattcac
      181 gctgaagcct acgctgctgg cgaactgaaa cacggtccgc tggcgctaat tgatgccgat
      241 atgccggtta ttgttgttgc accgaacaac gaattgctgg aaaaactgaa atccaacatt
      301 gaagaagttc gcgcgcgtgg cggtcagttg tatgtcttcg ccgatcagga tgcgggtttt
      361 gtaagtagcg ataacatgca catcatcgag atgccgcatg tggaagaggt gattgcaccg
      421 atcttctaca ccgttccgct gcagctgctg gcttaccatg tcgcgctgat caaaggcacc
      481 gacgttgacc agccgcgtaa cctggcaaaa tcggttacgg ttgagtaata aatggatctc
      541 ggtaccaaat tccagaaaag agacgctttc gagcgtcttt tttcgttttg gtccaccgta
      601 ttcatcagca ctgtcctgct ccttgtgatg gtttacaaac gtaaagagtc tctttaatac
      661 ctgtttttgc ttcatattgt tcagcgacag cttgctgtac ggcaggcacc agctcttccg
      721 ggatcagcgc gacgatacag ccgccaaatc cgccgccggt catgcgtacg ccacctttgt
      781 cgccaatcac agctttgacg atttctacca gagtgtcaat ttgcggcacg gtgatttcga
      841 aatcatcgcg catagaggca tgagactccg ccatcaactc gcccatacgt ttcaggtcgc
      901 cttgctccag cgcgctggca gcttcaacgg tgcgggcgtt ttcagtcagt atatgacgca
      961 cgcgttttgc cacgatcggg tccagttcat gcgcaacagc gttgaactct tcaatggtga
     1021 catcacgcag ggctggctgc tggaagaaac gcgcaccggt ttcgcactgt tcacgacggg
     1081 tgttgtattc gctgccaacc agggtacgtt tgaagttact gttgatgatg acgacagcca
     1141 cacctttggg catggaaact gctttggtcc ccagtgagcg gcaatcgatc agcaaggcat
     1201 gatctttctt gccgagcgcg gaaattagct gatccatgat cccgcagtta cagcctacaa
     1261 actggttttc tgcttcctga ccgttaagcg cgatttgtgc gccgtccagc ggcagatgat
     1321 aaagctgctg caatacggtt ccgaccgcga cttccagtga agcggaagaa cttaacccgg
     1381 caccctgcgg cacattgccg ctgatcacca tgtccacgcc gccgaagctg ttgttacgca
     1441 gttgcagatg tttcaccacg ccacgaacgt agttagccca ttgatagttt tcatgtgcga
     1501 caatgggcgc atcgagggaa aactcgtcga gctgattttc ataatcggct gccatcacgc
     1561 gaactttacg gtcatcgcgt ggtgcacaac tgatcacggt ttgataatca atcgcgcagg
     1621 gcagaacgaa accgtcgttg tagtcggtgt gttcaccaat caaattcacg cggccaggcg
     1681 cctgaatggt gtgagtggca gggtagccaa atgcgttggc aaacagagat tgtgtttttt
     1741 ctttcagact catgattgcc tcctgggtta gttcctcacc ttgtcgtatt atactatgcc
     1801 gatatactat gccgatgatt aattgtcaac ctttaatgaa ttataaaacg aaaggctcag
     1861 tcgaaagact gggcctttcg ttttatgcgt tgcgcttaat ctttctgcga attgagatga
     1921 cgccactggc tgggcgtcat cccggtttcc cgggtaaaca ccaccgaaaa atagttacta
     1981 tcttcaaagc cacattcggt cgaaatatca ctgattaaca ggcggctatg ctggagaaga
     2041 tattgcgcat gacacactct gacctgtcgc agatattgat tgatggtcat tccagtctgc
     2101 tggcgaaatt gctgacgcaa aacgcgctca ctgcacgatg cctcatcaca aaatttatcc
     2161 agcgcaaagg gacttttcag gctagccgcc agccgggtaa tcagcttatc cagcaacgtt
     2221 tcgctggatg ttggcggcaa cgaatcactg gtgtaacgat ggcgattcag caacatcacc
     2281 aactgcccga acagcaactc agccatttcg ttagcaaacg gcacatgctg actactttca
     2341 tgctcaagct gaccaataac ctgccgcgcc tgcgccatcc ccatgctacc taagcgccag
     2401 tgtggttgcc ctgcgctggc gttaaatccc ggaatcgccc cctgccagtc aagattcagc
     2461 ttcagacgct ccgggcaata aataatattc tgcaaaacca gatcgttaac ggaagcgtag
     2521 gagtgtttat catcagcatg aatgtaaaag agatcgccac gggtaatgcg ataagggcga
     2581 tcgttgagta catgcaggcc attaccgcgc cagacaatca ccagctcaca aaaatcatgt
     2641 gtatgttcag caaagacatc ttgcggataa cggtcagcca cagcgactgc ctgctggtcg
     2701 ctggcaaaaa aatcatcttt gagaagtttt aactgatgcg ccaccgtggc tacctcggcc
     2761 agagaacgaa gttgattatt cgcaatatgg cgtacaaata cgttgagaag attcgcgtta
     2821 ttgcagaaag ccatcccgtc cctggcgaat atcacgcggt gaccagttaa actctcggcg
     2881 aaaaagcgtc gaaaagtggt tactgtcgct gaatccacag cgataggcga tgtcagtaac
     2941 gctggcctcg ctgtggcgta gcagatgtcg ggctttcatc agtcgcaggc ggttcaggta
     3001 tcgctgaggc gtcagtcccg tttgctgctt aagctgccga tgtagcgtac gcagtgaaag
     3061 agaaaattga tccgccacgg catcccaatt cacctcatcg gcaaaatggt cctccagcca
     3121 ggccagaagc aagttgagac gtgatgcgct gttttccagg ttctcctgca aactgctttt
     3181 acgcagcaag agcagtaatt gcataaacaa gatctcgcga ctggcggtcg agggtaaatc
     3241 attttcccct tcctgctgtt ccatctgtgc aaccagctgt cgcacctgct gcaatacgct
     3301 gtggttaacg cgccagtgag acggatactg cccatccagc tcttgtggca gcaactgatt
     3361 cagcccggcg agaaactgaa atcgatccgg cgagcgatac agcacattgg tcagacacag
     3421 attatcggta tgttcataca gatgccgatc atgatcgcgt acgaaacaga ccgtgccacc
     3481 ggtgatggta tagggctgcc cattaaacac atgaataccc gtgccatgtt cgacaatcac
     3541 aatttcatga aaatcatgat gatgttcagg aaaatccgcc tgcgggagcc ggggttctat
     3601 cgccacggac gcgttaccag acggaaaaaa atccacacta tgtaatacgg tcatactggc
     3661 ctcctgatgt cgtcaacacg gcgaaatagt aatcacgagg tcaggttctt accttaaatt
     3721 ttcgacggaa aaccacgtaa aaaacgtcga tttttcaaga tacagcgtga attttcagga
     3781 aatgcggtga gcatcacatc accacaattc agcaaattgt gaacatcatc acgttcatct
     3841 ttccctggtt gccaatggcc cattttcttg tcagtaacga gaaggtcgcg aatccaggcg
     3901 ctttttagac tggtcgtata ctagagatta aagaggagaa aggggggatg aaggctattt
     3961 tcgtactgaa acatcttaat catgctaagg aggttttcta atgagcaaag gagaagaact
     4021 tttcactgga gttgtgccaa ttcttgttga attagatggt gatgttaatg ggcacaaatt
     4081 ttctgtcagt ggagagggtg aaggtgatgc tacatacgga aaactcaccc ttaaatttat
     4141 ttgcactact ggaaaactac ctgttccatg gccaacactt gtcactactc tgacctatgg
     4201 cgttcaatgc ttttcccgtt atccggatca catgaaacgg catgactttt tcaagagtgc
     4261 catgcccgaa ggttatgtac aggaacgcac tatatctttc aaagatgacg ggaactacaa
     4321 gacgcgtgct gaagtcaagt ttgaaggtga tacccttgtt aatcgtatcg agttaaaggg
     4381 tattgatttt aaagaagatg gaaacattct cggacacaaa ctagagtaca actataactc
     4441 acacaatgta tacatcacgg cagacaaaca aaagaatgga atcaaagcta acttcaaaat
     4501 tcgccacaac attgaagatg gttccgttca actagcagac cattatcaac aaaatactcc
     4561 aattggcgat ggccctgtcc ttttaccaga caaccattac ctgtcgacac aatctgccct
     4621 ttcgaaagat cccaacgaaa agcgtgacca catggtcctt cttgagtttg taactgctgc
     4681 tgggattaca catggcatgg atgagctcta caaatgacca attattgaag gccgctaacg
     4741 cggccttttt ttgtttctgg tctcccaaaa atttatttgc ttattaatca tccggctcgt
     4801 ataatgtgtg gagggcccaa gttcacttaa aaaggagaat agttatgaaa tcgaacaatg
     4861 cgctcatcgt catcctcggc accgtcaccc tggatgctgt aggcataggc ttggttatgc
     4921 cggtactgcc gggcctcttg cgggatatcg tccattccga cagcatcgcc agtcactatg
     4981 gcgtgctgct agcgctatat gcgttgatgc aatttctatg cgcacccgtt ctcggagcac
     5041 tgtccgaccg ctttggccgc cgcccagtcc tgctcgcttc gctacttgga gccactatcg
     5101 actacgcgat catggcgacc acacccgtcc tgtggattct ctacgccgga cgcatcgtgg
     5161 ccggcatcac cggcgccaca ggtgcggttg ctggcgccta tatcgccgac atcaccgatg
     5221 gggaagatcg ggctcgccac ttcgggctca tgagcgcttg tttcggcgtg ggtatggtgg
     5281 caggccccgt ggccggggga ctgttgggcg ccatctccct gcacgcacca ttccttgcgg
     5341 cggcggtgct caacggcctc aacctactac tgggctgctt cctaatgcag gagtcgcata
     5401 agggagagcg ccgtccgatg cccttgagag ccttcaaccc agtcagctcc ttccggtggg
     5461 cgcggggcat gaccattgtg gccgcactta tgactgtctt ctttatcatg caactcgtag
     5521 gacaggtgcc ggcagcgctc tgggtcattt tcggcgagga ccgctttcgc tggagcgcga
     5581 cgatgagcgg cctgtcgctt gcggtattcg gaatcttgca cgccctcgct caagccttcg
     5641 tcactggtcc cgccaccaaa cgtttcggcg agaagcaggc cattatcgcc ggcatggcgg
     5701 ccgacgcgct gggctacgtc ttgctggcgt tcgcgacgcg aggctggatg gccttcccca
     5761 ttatgattct tctcgcttcc ggcggcatcg ggatgcccgc gttgcaggcc atgctgtcca
     5821 ggcaggtaga tgacgaccat cagggacagc ttcaaggatc gctcgcggct cttaccagcc
     5881 taacttcgat cactggaccg ctgatcgtca cggcgattta tgccgcctcg gcgagcacat
     5941 ggaacgggtt ggcatggatt gtaggcgccg ccctatacct tgtctgcctc cccgcgttgc
     6001 gtcgcggtgc atggagccgg gccacctcga cctgagacaa ttgtccagat aaaaaaaatc
     6061 cttagctttc gctaaggatg atttctaaaa tttttctgtc atctcttcgt tattaatgtt
     6121 tgtaattgac tgaatatcaa cgcttattta aatcagactg aagactttat ctctctgtca
     6181 taaaactgtc atattcctta catataactg tcacctgttt gtcctatttt gcttctcgta
     6241 gccaacaaac aatgctttat gaatcctccc aggagacatt atgaaagtta tgcgtaccac
     6301 cgtcgcaact gttgtcgccg cgaccttatc gatgagtgct ttctctgtgt ttgcagaagc
     6361 aagcctgaca ggtgcaggtg caaccttccc tgcgccggtg tatgccaaat gggctgacac
     6421 ttaccagaaa gaaaccggta ataaagttaa ctaccagggt atcggttctt ccggtggcgt
     6481 aaaacagatt atcgctaata ccgttgattt tggtgcctct gacgcgcc
//
