LOCUS       sequence-131026-        4898 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Acetobacter aceti 1023 citrate synthase (AaCSH6) in E.
            coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4898)
  AUTHORS   Francois JA, Starks CM, Sivanuntakorn S, Jiang H, Ransome AE, Nam 
            JW, Constantine CZ, Kappock TJ
  TITLE     Structure of a NADH-insensitive hexameric citrate synthase that 
            resists acid inactivation.
  JOURNAL   Biochemistry. 2006 Nov 14;45(45):13487-99.
  PUBMED    17087502
REFERENCE   2  (bases 1 to 4898)
  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: 
            "Biochemistry."; date: "2006-11-14"; volume: "45(45)"; pages: 
            "13487-99"
COMMENT     Sequence Label: pJK225
FEATURES             Location/Qualifiers
     source          1..4898
                     /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"
     RBS             1479..1501
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(1532..1551)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1533..1551)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2060..2251
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2245..2267)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2353..2495
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2408..2427
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2510..2527)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2681..3269)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2761..2780)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3440..4231,4232..4300))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4063..4082
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4301..4405)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4432..4887)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4569..4590)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4781..4800
                     /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 gaggcgtttg gcaagcggca
      181 catagtcacg ctgcggtgcg ccaatataaa gctggcgagg gcggctgata cgctggcccg
      241 gttcttcaat catttccttc cactggctca cccagccggt ggtgcgggct acggcaaaca
      301 gcacagtaaa catactggtg gggatgccca tggccttgag aatgatgcca gagtagaaat
      361 ccacattcgg gtaaagtttg cgctgcacga agtaatcatc gctcagagca atcttttcca
      421 gctcaaccgc cagatccagc agcggatcat ccttaatgcc aagttctgtc agcacttcgt
      481 ggcaggtctg ctgcatgatc ttcgcacgtg ggtcgaagtt cttgtaaacg cggtggccaa
      541 agcccatcag ctttacgccg ctgttcttgt ccttcacctg tgcgataaag gcaggaatat
      601 tttctttctt gccaatacgg gccagcattt tcagcacagc ttcgtttgcg ccaccatgtg
      661 caggtcccca cagagcggca atgcccgcag caatacaggc aaacggattg gcccctgtag
      721 aaccagccag acgtacggtg gaggtagagg cattctgctc atgatcggca tgcagaatca
      781 gaatccggtt catggcgcgg gccagaacag ggttgacctt gtaaggttcg gacatgcgcg
      841 cgaacatcat ggacaggaag ttttctgcgt agttcagatc attccgcggg tagataaagg
      901 cttcaccctg cgtgtatttg taagcccatg ccgcaatggt tgggattttg gcaatcagcc
      961 gcatggcggc cagatcccga ttggcgggaa tggcaatatc gttggcatct gggtagaagg
     1021 cagacaaagc cccaaccgta ccacacagaa tggccattgg gtgggcatca cgccggaagc
     1081 cgttaaagaa gttacggatc tgctcgtgca gcagcgtatg gtttgtaagg gtgttggtga
     1141 aggtgtcgta ctgcgccttg ttgggcagtt cgccattcaa aagcagataa ataacttctt
     1201 cgtaggaagc attttcggcc agctgcgcaa tagggtaacc acggtgcagc agaacgcctt
     1261 tatcaccatc aataaaggtg attttgctgt tgcaggccgc tgtttccccg taacctgggt
     1321 cgaacgtgaa aacgcccagt tgcgccggaa gtttgcggat gtcgataaca tccggtccca
     1381 gagtgcctga aagcacaggc atttcggcgg attttccatc aaccgaaatg gtagcggtag
     1441 atagcttacc ttctttctgc gacgcgctca tatgtatatc tccttcttaa agttaaacaa
     1501 aattatttct agagggaaac cgttgtggtc tccctatagt gagtcgtatt aatttcgcgg
     1561 gatcgagatc tcgggcagcg ttgggtcctg gccacgggtg cgcatgatcg tgctcctgtc
     1621 gttgaggacc cggctaggct ggcggggttg ccttactggt tagcagaatg aatcaccgat
     1681 acgcgagcga acgtgaagcg actgctgctg caaaacgtct gcgacctgag caacaacatg
     1741 aatggtcttc ggtttccgtg tttcgtaaag tctggaaacg cggaagtcag cgccctgcac
     1801 cattatgttc cggatctgca tcgcaggatg ctgctggcta ccctgtggaa cacctacatc
     1861 tgtattaacg aagcgctggc attgaccctg agtgattttt ctctggtccc gccgcatcca
     1921 taccgccagt tgtttaccct cacaacgttc cagtaaccgg gcatgttcat catcagtaac
     1981 ccgtatcgtg agcatcctct ctcgtttcat cggtatcatt acccccatga acagaaatcc
     2041 cccttacacg gaggcatcag tgaccaaaca ggaaaaaacc gcccttaaca tggcccgctt
     2101 tatcagaagc cagacattaa cgcttctgga gaaactcaac gagctggacg cggatgaaca
     2161 ggcagacatc tgtgaatcgc ttcacgacca cgctgatgag ctttaccgca gctgcctcgc
     2221 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     2281 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     2341 cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat agcggagtgt atactggctt
     2401 aactatgcgg catcagagca gattgtactg agagtgcacc atatatgcgg tgtgaaatac
     2461 cgcacagatg cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg
     2521 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     2581 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     2641 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     2701 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     2761 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     2821 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     2881 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     2941 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     3001 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     3061 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     3121 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     3181 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     3241 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     3301 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     3361 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     3421 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     3481 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     3541 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     3601 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     3661 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     3721 cagttaatag tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg tcacgctcgt
     3781 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     3841 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     3901 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     3961 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     4021 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     4081 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     4141 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     4201 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     4261 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     4321 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     4381 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gaaattgtaa
     4441 acgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     4501 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     4561 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     4621 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     4681 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     4741 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     4801 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     4861 cgcttaatgc gccgctacag ggcgcgtccc attcgcca
//
