LOCUS       sequence-426101-        7697 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  amyE integration vector for IPTG-inducible expression in Bacillus 
            subtilis (Pspank-Hy promoter).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7697)
  TITLE     A historical sequence deletion in a commonly used Bacillus subtilis 
            chromosome integration vector generates undetected loss-of-function 
            mutations
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7697)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pGL004
FEATURES             Location/Qualifiers
     source          1..7697
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(2825..3413)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2905..2924)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3584..4375,4376..4444))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4207..4226
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4445..4549)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4617..4635
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             complement(5449..6531)
                     /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
                     EACKTAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     6493..6512
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    6803..6819
                     /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)."
     terminator      complement(6923..6957)
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     terminator      complement(7037..7064)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(7083..7169)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 aaaaaaaata ggagataaaa gcaacttacg atatattgaa ttaacaatta ttattcagca
       61 agaaatggta ccgtggaatc atcctcccaa acaagaattt atttatggag aatggttaca
      121 agagctttat gaacaaggat acattcctca gaaggaatta aattcagatt taaccataat
      181 gctttaccaa gcaaaacgaa aaaataaaag aatatacgga aattatgact tagaggaatt
      241 actacctgat attccatttt ctgatgtgag aagagccatt atggattcgt cagaggaatt
      301 aatagataat tatcaggatg atgaaaccaa ctctatatta actttatgcc gtatgatttt
      361 aactatggac acgggtaaaa tcataccaaa agatattgcg ggaaatgcag tggctgaatc
      421 ttctccatta gaacataggg agagaatttt gttagcagtt cgtagttatc ttggagagaa
      481 tattgaatgg actaatgaaa atgtaaattt aactataaac tatttaaata acagattaaa
      541 aaaattataa aaaaattgaa aaaatggtgg aaacactttt ttcaattttt ttgttttatt
      601 atttaatatt tgggaaatat tcattctaat tggtaatcag attttagaaa acaataaacc
      661 cttgcatagg gggaatccgt ttaggctggg cggtgatagc ttctcgttca ggcagtacgc
      721 ctcttttctt ttccagacct gagggaggcg gaaatggtgt gaggttcccg gggaaaagcc
      781 aaataggcga tcgcgggagt gctttatttg aagatcaggc tatcactgcg gtcaatagat
      841 ttcacaatgt gatggctgga cagcctgagg aactctcgaa cccgaatgga aacaaccaga
      901 tatttatgaa tcagcgcggc tcacatggcg ttgtgctggc aaatgcaggt tcatcctctg
      961 tctctatcaa tacggcaaca aaattgcctg atggcaggta tgacaataaa gctggagcgg
     1021 gttcatttca agtgaacgat ggtaaactga caggcacgat caatgccagg tctgtagctg
     1081 tgctttatcc tgatgatatt gcaaaagcgc ctcatgtttt ccttgagaat tacaaaacag
     1141 gtgtaacaca ttctttcaat gatcaactga cgattacctt gcgtgcagat gcgaatacaa
     1201 caaaagccgt ttatcaaatc aataatggac cagagacggc gtttaaggat ggagatcaat
     1261 tcacaatcgg aaaaggagat ccatttggca aaacatacac catcatgtta aaaggaacga
     1321 acagtgatgg tgtaacgagg accgagaaat acagttttgt taaaagagat ccagcgtcgg
     1381 ccaaaaccat cggctatcaa aatccgaatc attggagcca ggtaaatgct tatatctata
     1441 aacatgatgg gagccgagta attgaattga ccggatcttg gcctggaaaa ccaatgacta
     1501 aaaatgcaga cggaatttac acgctgacgc tgcctgcgga cacggataca accaacgcaa
     1561 aagtgatttt taataatggc agcgcccaag tgcccggtca gaatcagcct ggctttgatt
     1621 acgtgctaaa tggtttatat aatgactcgg gcttaagcgg ttctcttccc cattgagggc
     1681 aaggctagac gggacttacc gaaagaaacc atcaatgatg gtttcttttt tgttcataaa
     1741 tcagacaaaa cttttctctt gcaaaagttt gtgaagtgtt gcacaatata aatgtgaaat
     1801 acttcacaaa caaaaagaca tcaaagagaa acataccgcg aggctggatg gccttcccca
     1861 ttatgattct tctcgcttcc ggcggcatcg ggatgcccgc gttgcaggcc atgctgtcca
     1921 ggcaggtaga tgacgaccat cagggacagc ttcaaggatc gctcgcggct cttaccagcc
     1981 taacttcgat cattggaccg ctgatcgtca cggcgattta tgccgcctcg gcgagcacat
     2041 ggaacgggtt ggcatggatt gtaggcgccg ccctatacct tgtctgcctc cccgcgttgc
     2101 gtcgcggtgc atggagccgg gccacctact gaagtggatt tctttaagag ctcctttaac
     2161 ttcctcacca gtagttgtat cggtaccata agtagaagca gcaacccaag tagctttacc
     2221 agcatccggt tcaaccagca tagtaagaat cttactggac atcggcagtt cttcgaacag
     2281 tgcgccaact accagctctt tctgcagttc attcagggca ccggagaacc tgcgtgcaat
     2341 ccatcttgtt caatcatgcg aaacgatcct catcctgtct cttgatccat ggattacgcg
     2401 ttaacccggg cccgcggatg catatgatca gatcttaagg cctaggtcta gagtctttgt
     2461 tttgacgcca ttagcgtacg taacaatcct cgttaaagga caaggacctg agcggaagtg
     2521 tatcgtacag tagacggagt atactagtat agtctatagt ccgtggaatt attatattta
     2581 tctccgacga tattctcatc agtgaaatcc agctggagtt ctttagcaaa tttttttatt
     2641 agctgaactt agtattagtg gccatactcc tccaatccaa agctatttag aaagattact
     2701 atatcctcaa acaggcggta accggcctct tcatcgggaa tgcgcgcgac cttcagcatc
     2761 gccggcatgt ccccctggcg gacgggaagt atccagctcg aggtcgggcc gcgttgctgg
     2821 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     2881 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     2941 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     3001 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     3061 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     3121 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     3181 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     3241 ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag
     3301 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     3361 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     3421 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     3481 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     3541 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     3601 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     3661 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     3721 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     3781 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     3841 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgctg
     3901 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     3961 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     4021 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     4081 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     4141 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     4201 cacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     4261 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     4321 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     4381 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     4441 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     4501 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     4561 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
     4621 ggcgtatcac gaggcccttt cgtcttcaag aattaacaaa attctccagt cttcacatcg
     4681 gtttgaaagg aggaagcgga agaatgaagt aagagggatt tttgactccg aagtaagtct
     4741 tcaaaaaatc aaataaggag tgtcaagaat gtttgcaaaa cgattcaaaa cctctttact
     4801 gccgttattc gctggatttt tattgctgtt tcatttggtt ctggcaggac cggcggctgc
     4861 gagtgctgaa acggcgaaca aatcgaatga gcttacagca ccgtcgatca aaagcggaac
     4921 cattcttcat gcatggaatt ggtcgttcaa tacgttaaaa cacaatatga aggatattca
     4981 tgatgcagga tatacagcca ttcagacatc tccgattaac caagtaaagg aagggaatca
     5041 aggagataaa agcatgtcga actggtactg gctgtatcag ccgacatcgt atcaaattgg
     5101 caaccgttac ttaggtactg aacaagaatt taaagaaatg tgtgcagccg ctgaagaata
     5161 tggcataaag gtcattgttg acgcggtcat caatcatacc accagtgatt atgccgcgat
     5221 ttccaatgag gttaagagta ttccaaactg gacacatgga aacacacaaa ttaaaaactg
     5281 gtctgatcga tgggatgtca cgcagaatta aatagtacat aatggatttc cttacgcgaa
     5341 atacgggcag acatggcctg cccggttatt attatttttg acaccagacc aactggtaat
     5401 ggtagcgacc ggcgctcagg atcctaactc acattaattg cgttgcgctc actgcccgct
     5461 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     5521 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag
     5581 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg
     5641 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc
     5701 ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt
     5761 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac
     5821 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc
     5881 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag
     5941 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc
     6001 caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact
     6061 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc
     6121 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg
     6181 ttgcgcgaga agattgtgca ccgccgtttt acaggcttcg acgccgcttc gttctaccat
     6241 cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg
     6301 cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc
     6361 cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac
     6421 tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg
     6481 ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca tcaaaatcgt
     6541 ctccctccgt ttgaatattt gattgatcgt aaccagatga agcactcttt ccactatccc
     6601 tacagtgtta tggcttgaac aatcacgaaa caataattgg tacgtacgat ctttcagccg
     6661 actcaaacat caaatcttac aaatgtagtc tttgaaagta ttacatatgt aagatttaaa
     6721 tgcaaccgtt ttttcggaag gaaatgatga cctcgtttcc accgaattag cttgcatgcg
     6781 gctagctgtc gactaagctt aattgttatc cgctcacaat tacacacatt atgccacacc
     6841 ttgtagataa agtcaacaac ttttgcaaaa tgaattgtga gtgctcacat ttaccctcga
     6901 gcaacgttct tgccattgct gcataaaaaa cgcccggcgg caaccgagcg ttctgaatta
     6961 attaatcatc gggaagatct tcatcaccga aacgcggcag gcagctctag agttaacaag
     7021 agtttgtaga aacgcaaaaa ggccatccgt caggatggcc ttctgcttag ctagagcggc
     7081 ggatttgtcc tactcaggag agcgttcacc gacaaacaac agataaaacg aaaggcccag
     7141 tctttcgact gagcctttcg ttttatttga tgcctcaagc tagaggtcga attcctgcag
     7201 ctggcgaatg gcgattttcg ttcgtgaata catgttataa taactataac taataacgta
     7261 acgtgactgg caagagatat ttttaaaaca atgaataggt ttacacttac tttagtttta
     7321 tggaaatgaa agatcatatc atatataatc tagaataaaa ttaactaaaa taattattat
     7381 ctagataaaa aatttagaag ccaatgaaat ctataaataa actaaattaa gtttatttaa
     7441 ttaacaacta tggatataaa ataggtacta atcaaaatag tgaggaggat atatttgaat
     7501 acatacgaac aaattaataa agtgaaaaaa atacttcgga aacatttaaa aaataacctt
     7561 attggtactt acatgtttgg atcaggagtt gagagtggac taaaaccaaa tagtgatctt
     7621 gactttttag tcgtcgtatc tgaaccattg acagatcaaa gtaaagaaat acttatacaa
     7681 aaaattagac ctatttc
//
