LOCUS       sequence-232493-        5868 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  precursor slow-folding maltodextrin binding protein, MBP or malE, 
            with mutations Y283D, G289C and V347C.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5868)
  AUTHORS   Bariya P, Randall LL
  TITLE     Co-assembly of SecYEG and SecA fully restores the properties of the 
            native translocon.
  JOURNAL   J Bacteriol. 2018 Oct 1. pii: JB.00493-18. doi: 10.1128/JB.00493-18.
  PUBMED    30275279
REFERENCE   2  (bases 1 to 5868)
  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"; journalName: "J 
            Bacteriol. 2018 Oct 1. pii: JB.00493-18. doi: 10.1128/JB.00493-18."
COMMENT     Sequence Label: pAL829
FEATURES             Location/Qualifiers
     source          1..5868
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     3..21
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(45..59,60..65,66..82,83..89)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    65..81
                     /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)."
     protein_bind    97..113
                     /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)."
     RBS             132..143
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             join(150..227,228..1325)
                     /codon_start=1
                     /gene="malE"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This gene encodes a signal sequence that can direct 
                     MBP to the bacterial periplasm."
                     /translation="MKIKTGARILALSALTTMMFSASALAKIEEGKLVIWINGDKGYNG
                     LAEVGKKFEKDTGIKVTVEHPDKLEEKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAE
                     ITPDKAFQDKLYPFTWDAVRYNGKLIAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDK
                     ELKAKGKSALMFNLQEPYFTWPLIAADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLV
                     DLIKNKHMNADTDYSIAEAAFNKGETAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPS
                     KPFVGVLSAGINAASPNKELAKEFLENDLLTDECLEAVNKDKPLGAVALKSYEEELAKD
                     PRIAATMENAQKGEIMPNIPQMSAFWYAVRTACINAASGRQTVDEALKDAQT"
     regulatory      1182..1191
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1299..1322
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     terminator      1360..1454
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1498..2157
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1564..1583)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2222..2308
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2401..3483)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     3445..3464
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3484..3561)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     complement(3645..3667)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3753..3893
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3808..3827
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3908..3925)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4079..4667)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4159..4178)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4838..5629,5630..5698))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5461..5480
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5699..5803)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaaataggcg tatcacgagg ccctttcgtc ttcacctcga gaaatcataa aaaatttatt
       61 tgctttgtga gcggataaca attataatag attcaattgt gagcggataa caatttcaca
      121 cagaattcat taaagaggag aaattaacta tgaaaataaa aacaggtgca cgcatcctcg
      181 cattatccgc attaacgacg atgatgtttt ccgcctcggc tctcgccaaa atcgaagaag
      241 gtaaactggt aatctggatt aacggcgata aaggctataa cggtctcgct gaagtcggta
      301 agaaattcga gaaagatacc ggaattaaag tcaccgttga gcatccggat aaactggaag
      361 agaaattccc acaggttgcg gcaactggcg atggccctga cattatcttc tgggcacacg
      421 accgctttgg tggctacgct caatctggcc tgttggctga aatcaccccg gacaaagcgt
      481 tccaggacaa gctgtatccg tttacctggg atgccgtacg ttacaacggc aagctgattg
      541 cttacccgat cgctgttgaa gcgttatcgc tgatttataa caaagatctg ctgccgaacc
      601 cgccaaaaac ctgggaagag atcccggcgc tggataaaga actgaaagcg aaaggtaaga
      661 gcgcgctgat gttcaacctg caagaaccgt acttcacctg gccgctgatt gctgctgacg
      721 ggggttatgc gttcaagtat gaaaacggca agtacgacat taaagacgtg ggcgtggata
      781 acgctggcgc gaaagcgggt ctgaccttcc tggttgacct gattaaaaac aaacacatga
      841 atgcagacac cgattactcc atcgcagaag ctgcctttaa taaaggcgaa acagcgatga
      901 ccatcaacgg cccgtgggca tggtccaaca tcgacaccag caaagtgaat tatggtgtaa
      961 cggtactgcc gaccttcaag ggtcaaccat ccaaaccgtt cgttggcgtg ctgagcgcag
     1021 gtattaacgc cgccagtccg aacaaagagc tggcgaaaga gttcctcgaa aacgatctgc
     1081 tgactgacga atgtctggaa gcggttaata aagacaaacc gctgggtgcc gtagcgctga
     1141 agtcttacga ggaagagttg gcgaaagatc cacgtattgc cgccaccatg gaaaacgccc
     1201 agaaaggtga aatcatgccg aacatcccgc agatgtccgc tttctggtat gccgtgcgta
     1261 ctgcgtgtat caacgccgcc agcggtcgtc agactgtcga tgaagccctg aaagacgcgc
     1321 agactcgtat caccaagtaa gcttaattag ctgagcttgg actcctgttg atagatccag
     1381 taatgacctc agaactccat ctggatttgt tcagaacgct cggttgccgc cgggcgtttt
     1441 ttattggtga gaatccaagc tagcttggcg agattttcag gagctaagga agctaaaatg
     1501 gagaaaaaaa tcactggata taccaccgtt gatatatccc aatggcatcg taaagaacat
     1561 tttgaggcat ttcagtcagt tgctcaatgt acctataacc agaccgttca gctggatatt
     1621 acggcctttt taaagaccgt aaagaaaaat aagcacaagt tttatccggc ctttattcac
     1681 attcttgccc gcctgatgaa tgctcatccg gaatttcgta tggcaatgaa agacggtgag
     1741 ctggtgatat gggatagtgt tcacccttgt tacaccgttt tccatgagca aactgaaacg
     1801 ttttcatcgc tctggagtga ataccacgac gatttccggc agtttctaca catatattcg
     1861 caagatgtgg cgtgttacgg tgaaaacctg gcctatttcc ctaaagggtt tattgagaat
     1921 atgtttttcg tctcagccaa tccctgggtg agtttcacca gttttgattt aaacgtggcc
     1981 aatatggaca acttcttcgc ccccgttttc accatgggca aatattatac gcaaggcgac
     2041 aaggtgctga tgccgctggc gattcaggtt catcatgccg tttgtgatgg cttccatgtc
     2101 ggcagaatgc ttaatgaatt acaacagtac tgcgatgagt ggcagggcgg ggcgtaattt
     2161 ttttaaggca gttattggtg cccttaaacg cctggggtaa tgactctcta gcttgaggca
     2221 tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt cgttttatct gttgtttgtc
     2281 ggtgaacgct ctcctgagta ggacaaatcc gccctctaga ttacgtgcag tcgatgataa
     2341 gctgtcaaac atgagaattg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc
     2401 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     2461 cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga
     2521 gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc
     2581 cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata
     2641 acatgagctg tcttcggtat cgtcgtatcc cactaccgag atatccgcac caacgcgcag
     2701 cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat
     2761 cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc
     2821 actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg
     2881 ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat
     2941 ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga
     3001 gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt
     3061 agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag
     3121 cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct
     3181 tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc
     3241 cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa
     3301 cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat
     3361 cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg
     3421 ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt
     3481 cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt
     3541 tttgcaccat tcgatggtgt cggaatttcg ggcagcgttg ggtcctggcc acgggtgcgc
     3601 atgatctaga gctgcctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag
     3661 ctcccggaga cggtcacagc ttgtctgtaa gcggatgacg ggagcagaca agcccgtcag
     3721 ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat
     3781 agcggagtgt atactggctt aactatgcgg catcagagca gattgtactg agagtgcacc
     3841 acatgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgctctt
     3901 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     3961 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     4021 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     4081 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     4141 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     4201 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     4261 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     4321 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     4381 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4441 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     4501 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4561 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     4621 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     4681 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     4741 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     4801 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     4861 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     4921 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     4981 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     5041 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     5101 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     5161 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     5221 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     5281 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     5341 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     5401 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     5461 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     5521 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     5581 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     5641 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     5701 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     5761 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     5821 tgccacctga cgtctaagaa accattatta tcatgacatt aacctata
//
