LOCUS       sequence-133863-        5514 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Produces Acetobacter aceti 1023 small hypothetical protein adjacent 
            to purEK genes, with N-terminal His6 tag (H6AaOrfX).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5514)
  AUTHORS   Kappock
  TITLE     Additional purEK associated constructs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5514)
  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            "Unpublished"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: pJK523
FEATURES             Location/Qualifiers
     source          1..5514
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(388..405)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(415..432)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             451..473
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    488..512
                     /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)."
     primer_bind     complement(512..531)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(513..531)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(615..634)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        840..917
                     /gene="lacI"
                     /label=lacI promoter
     CDS             918..2000
                     /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     complement(937..956)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             2809..3000
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2994..3016)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3102..3244
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3157..3176
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3259..3276)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3430..4018)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3510..3529)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4129..4148)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4140..4955
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             4140..4955
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4213..4232)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5048..5503)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5185..5206)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5397..5416
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttca
      181 cgcgatgtca tccggaatca gtagactttc cactttcgta atttgatcct taagcagcag
      241 tttgcgtttt ttcagacgtt gcagatacag ctgatcgata aacgtggttt catgggccat
      301 acggttgatc acggtgtcca gatcgcggtg ttcgctacgc agttcatgca attgcgccag
      361 cagcaggtcg cgatcacgca gcatatggct gccgcgcggc accaggccgc tgctgtgatg
      421 atgatgatga tggctgctgc ccatggtata tctccttctt aaagttaaac aaaattattt
      481 ctagagggga attgttatcc gctcacaatt cccctatagt gagtcgtatt aatttcgcgg
      541 gatcgagatc tcgatcctct acgccggacg catcgtggcc ggcatcaccg gcgccacagg
      601 tgcggttgct ggcgcctata tcgccgacat caccgatggg gaagatcggg ctcgccactt
      661 cgggctcatg agcgcttgtt tcggcgtggg tatggtggca ggccccgtgg ccgggggact
      721 gttgggcgcc atctccttgc atgcaccatt ccttgcggcg gcggtgctca acggcctcaa
      781 cctactactg ggctgcttcc taatgcagga gtcgcataag ggagagcgtc gagatcccgg
      841 acaccatcga atggcgcaaa acctttcgcg gtatggcatg atagcgcccg gaagagagtc
      901 aattcagggt ggtgaatgtg aaaccagtaa cgttatacga tgtcgcagag tatgccggtg
      961 tctcttatca gaccgtttcc cgcgtggtga accaggccag ccacgtttct gcgaaaacgc
     1021 gggaaaaagt ggaagcggcg atggcggagc tgaattacat tcccaaccgc gtggcacaac
     1081 aactggcggg caaacagtcg ttgctgattg gcgttgccac ctccagtctg gccctgcacg
     1141 cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg
     1201 tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc
     1261 tcgcgcaacg cgtcagtggg ctgatcatta actatccgct ggatgaccag gatgccattg
     1321 ctgtggaagc tgcctgcact aatgttccgg cgttatttct tgatgtctct gaccagacac
     1381 ccatcaacag tattattttc tcccatgaag acggtacgcg actgggcgtg gagcatctgg
     1441 tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc attaagttct gtctcggcgc
     1501 gtctgcgtct ggctggctgg cataaatatc tcactcgcaa tcaaattcag ccgatagcgg
     1561 aacgggaagg cgactggagt gccatgtccg gttttcaaca aaccatgcaa atgctgaatg
     1621 agggcatcgt tcccactgcg atgctggttg ccaacgatca gatggcgctg ggcgcaatgc
     1681 gcgccattac cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg
     1741 ataccgaaga cagctcatgt tatatcccgc cgttaaccac catcaaacag gattttcgcc
     1801 tgctggggca aaccagcgtg gaccgcttgc tgcaactctc tcagggccag gcggtgaagg
     1861 gcaatcagct gttgcccgtc tcactggtga aaagaaaaac caccctggcg cccaatacgc
     1921 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     1981 gactggaaag cgggcagtga gcgcaacgca attaatgtaa gttagctcac tcattaggca
     2041 ccgggatctc gaccgatgcc cttgagagcc ttcaacccag tcagctcctt ccggtgggcg
     2101 cggggcatga ctatcgtcgc cgcacttatg actgtcttct ttatcatgca actcgtagga
     2161 caggtgccgg cagcgctctg ggtcattttc ggcgaggacc gctttcgctg gagcgcgacg
     2221 atgatcggcc tgtcgcttgc ggtattcgga atcttgcacg ccctcgctca agccttcgtc
     2281 actggtcccg ccaccaaacg tttcggcgag aagcaggcca ttatcgccgg catggcggcc
     2341 ccacgggtgc gcatgatcgt gctcctgtcg ttgaggaccc ggctaggctg gcggggttgc
     2401 cttactggtt agcagaatga atcaccgata cgcgagcgaa cgtgaagcga ctgctgctgc
     2461 aaaacgtctg cgacctgagc aacaacatga atggtcttcg gtttccgtgt ttcgtaaagt
     2521 ctggaaacgc ggaagtcagc gccctgcacc attatgttcc ggatctgcat cgcaggatgc
     2581 tgctggctac cctgtggaac acctacatct gtattaacga agcgctggca ttgaccctga
     2641 gtgatttttc tctggtcccg ccgcatccat accgccagtt gtttaccctc acaacgttcc
     2701 agtaaccggg catgttcatc atcagtaacc cgtatcgtga gcatcctctc tcgtttcatc
     2761 ggtatcatta cccccatgaa cagaaatccc ccttacacgg aggcatcagt gaccaaacag
     2821 gaaaaaaccg cccttaacat ggcccgcttt atcagaagcc agacattaac gcttctggag
     2881 aaactcaacg agctggacgc ggatgaacag gcagacatct gtgaatcgct tcacgaccac
     2941 gctgatgagc tttaccgcag ctgcctcgcg cgtttcggtg atgacggtga aaacctctga
     3001 cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa
     3061 gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat gacccagtca
     3121 cgtagcgata gcggagtgta tactggctta actatgcggc atcagagcag attgtactga
     3181 gagtgcacca tatatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     3241 caggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     3301 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     3361 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     3421 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     3481 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     3541 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     3601 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     3661 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     3721 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     3781 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     3841 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa
     3901 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     3961 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4021 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4081 gattttggtc atgaacaata aaactgtctg cttacataaa cagtaataca aggggtgtta
     4141 tgagccatat tcaacgggaa acgtcttgct ctaggccgcg attaaattcc aacatggatg
     4201 ctgatttata tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct
     4261 atcgattgta tgggaagccc gatgcgccag agttgtttct gaaacatggc aaaggtagcg
     4321 ttgccaatga tgttacagat gagatggtca gactaaactg gctgacggaa tttatgcctc
     4381 ttccgaccat caagcatttt atccgtactc ctgatgatgc atggttactc accactgcga
     4441 tccccgggaa aacagcattc caggtattag aagaatatcc tgattcaggt gaaaatattg
     4501 ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt
     4561 ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg
     4621 ttgatgcgag tgattttgat gacgagcgta atggctggcc tgttgaacaa gtctggaaag
     4681 aaatgcataa acttttgcca ttctcaccgg attcagtcgt cactcatggt gatttctcac
     4741 ttgataacct tatttttgac gaggggaaat taataggttg tattgatgtt ggacgagtcg
     4801 gaatcgcaga ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc
     4861 cttcattaca gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat
     4921 tgcagtttca tttgatgctc gatgagtttt tctaagaatt aattcatgag cggatacata
     4981 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     5041 ccacctgaaa ttgtaaacgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     5101 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     5161 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     5221 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     5281 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     5341 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     5401 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     5461 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gcca
//
