LOCUS       sequence-308166-        6503 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6503)
  AUTHORS   Ha JY, Lee JH, Kim KH, Kim do J, Lee HH, Kim HK, Yoon HJ, Suh SW
  TITLE     Crystal Structure of d-Erythronate-4-phosphate Dehydrogenase 
            Complexed with NAD.
  JOURNAL   J Mol Biol. 2007 Mar 2. 366(4):1294-304.
  PUBMED    17217963
REFERENCE   2  (bases 1 to 6503)
  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 Mol Biol.
            2007 Mar 2. 366(4):1294-304."
COMMENT     Sequence Label: PA1375_pET21a(+)C-tag
FEATURES             Location/Qualifiers
     source          1..6503
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     171..190
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     274..293
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        274..292
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    293..317
                     /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             332..354
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1508..1525
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1578..1596)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1592..1639
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1676..2131
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1763..1782)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1973..1994
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2157..2261
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2262..2330,2331..3122)
                     /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     complement(2480..2499)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3293..3881
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3782..3801
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4035..4052
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4067..4206
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4132..4151)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4292..4314
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4308..4499)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5308..6390)
                     /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     6352..6371
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6391..6468)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 attaggaagc agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt
       61 gcatgcaagg agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg
      121 ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg
      181 gcgatatagg cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg
      241 gcgtagagga tcgagatctc gatcccgcga aattaatacg actcactata ggggaattgt
      301 gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag gagatataca
      361 tatgcgtatt ctcgccgatg aaaacattcc cgtggtcgac gccttcttcg ccgaccaggg
      421 ctccattcgc cgcttgcccg ggcgcgccat cgaccgcgcg gcgctggccg aggtcgacgt
      481 gctgctggtg cgctcggtca ccgaggtcag ccgcgcggcg ctggccggct cgccggtgcg
      541 cttcgtcggc acctgcacca tcggcaccga ccacctcgac ctcgactact tcgccgaggc
      601 cggcatcgcc tggtccagcg cacccggctg caacgcccgc ggggtggtgg actacgtgct
      661 cggctgcctg ctggcgatgg ccgaggtgcg cggcgccgac ctggccgaac gcacctatgg
      721 ggtggtcggc gccgggcagg tcggcgggcg gctggtggag gtgctgcgcg ggctgggctg
      781 gaaagtgctg gtctgcgatc cgccgcgcca ggcacgcgag ccggacggcg aattcgtctc
      841 gctggaacgg ctgctggccg aggccgacgt gatcagcctg cacactccgc tgaaccgcga
      901 cggcgaacac ccgacccgcc acctgctgga tgagccccgc ctggccgccc tgcgtcccgg
      961 cacctggctg gtcaacgcca gccgtggcgc ggtggtcgac aaccaggccc tgcgccgcct
     1021 gctcgaaggc ggcgccgacc tggaagtggc gctggacgtc tgggaaggcg aaccgcaggc
     1081 cgaccccgaa ctggccgcgc gctgcctgat cgccacgccg cacatcgccg gctacagcct
     1141 ggaaggcaag ctgcgcggca ccgcacagat ctaccaggcc tactgcgcct ggcgcggcat
     1201 cgccgaacgc gtgagcctgc aagacgtgct gccggaaacc tggctcgccg gcctgcaact
     1261 caaccccggc tgcgacccgg cctgggcgct ggccacgctg tgccgcgcgg tgtacgaccc
     1321 gcgcagcgac gacgccgcct tccgccgcag cctgaccggc gacagcgcca cccgccgcgc
     1381 cgccttcgac gccctgcgca aacactaccc gccgcgccgc gagatcaccg gcctgcgcgt
     1441 cgccaccggt ggacaggcgg agttgcagcg ggtggtgcgg gcgctggggg cgcagttggt
     1501 cctcgagcac caccaccacc accactgaga tccggctgct aacaaagccc gaaaggaagc
     1561 tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg
     1621 ggtcttgagg ggttttttgc tgaaaggagg aactatatcc ggattggcga atgggacgcg
     1681 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     1741 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     1801 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     1861 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     1921 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     1981 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     2041 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     2101 aattttaaca aaatattaac gcttacaatt taggtggcac ttttcgggga aatgtgcgcg
     2161 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     2221 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     2281 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     2341 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     2401 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     2461 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     2521 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     2581 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     2641 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     2701 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     2761 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgcagca atggcaacaa
     2821 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     2881 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     2941 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     3001 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     3061 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     3121 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     3181 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     3241 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     3301 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     3361 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     3421 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     3481 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     3541 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     3601 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     3661 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     3721 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     3781 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     3841 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     3901 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     3961 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     4021 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt
     4081 ttctccttac gcatctgtgc ggtatttcac accgcaatgg tgcactctca gtacaatctg
     4141 ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg
     4201 gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     4261 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     4321 ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc
     4381 gattcacaga tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt
     4441 aatgtctggc ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact
     4501 gatgcctccg tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag
     4561 aggatgctca cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag
     4621 ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc
     4681 cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg
     4741 cagatccgga acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca
     4801 cggaaaccga agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg
     4861 cttcacgttc gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag
     4921 cctagccggg tcctcaacga caggagcacg atcatgcgca cccgtggggc cgccatgccg
     4981 gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga
     5041 gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt cgcgctccag
     5101 cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc tacgagttgc
     5161 atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc ccaccggaag
     5221 gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc ctaatgagtg
     5281 agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     5341 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     5401 cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg
     5461 gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg
     5521 tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac
     5581 taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag
     5641 cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg
     5701 catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg
     5761 aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga
     5821 acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac
     5881 gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga
     5941 gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg
     6001 gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac
     6061 cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc
     6121 cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag
     6181 actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg
     6241 gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga
     6301 aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc
     6361 tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg
     6421 gcgctatcat gccataccgc gaaaggtttt gcgccattcg atggtgtccg ggatctcgac
     6481 gctctccctt atgcgactcc tgc
//
