LOCUS       sequence-130092-        5857 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 5857)
  AUTHORS   Kim KK, Yokota H, Santoso S, Lerner D, Kim R, Kim SH
  TITLE     Purification, crystallization, and preliminary X-ray 
            crystallographic data analysis of small heat shock protein homolog 
            from Methanococcus jannaschii, a hyperthermophile.
  JOURNAL   J Struct Biol. 1998 Jan . 121(1):76-80.
  PUBMED    9573624
REFERENCE   2  (bases 1 to 5857)
  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 Struct 
            Biol. 1998 Jan . 121(1):76-80."
COMMENT     Sequence Label: pET21a.MJ0285
FEATURES             Location/Qualifiers
     source          1..5857
                     /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             661..683
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    698..722
                     /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(722..741)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(723..741)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(825..844)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1050..1127
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1128..2210
                     /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(1147..1166)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3019..3210
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3204..3226)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3312..3454
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3367..3386
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3469..3486)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3640..4228)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3720..3739)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4399..5190,5191..5259))
                     /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     5022..5041
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5260..5364)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5391..5846)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5528..5549)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5740..5759
                     /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 gcaagcttgt
      181 cgacggagct cgaattcgga tccctcgagt cattcaatgt tgattccttt cttaattgag
      241 gattctgcct ttggtaatat aactgataaa acaccatttt caaacttagc tgaggcattt
      301 tcttccttaa cagttgcagg aagttttatt gttctatata tttcttcctc ttctggaatt
      361 tctgagtaga taattctttc actctcagtt atcattaatg ggcttctctt agctctaatc
      421 tctaatgtat ctccaactgc atttaaaatt atgtcctctt tattaacccc tggcaaccat
      481 gcaataactt ttatatgctg gtctccttca ataattgaga ttggcatgaa cccttttcca
      541 gaaatttgta ttcctgttga gctttgaatc attgtggttc ctgtcattgg tgttgcaaaa
      601 aactctttaa acattctttc aaataatgaa tcaaatgggt ctcttccgaa catatgtata
      661 tctccttctt aaagttaaac aaaattattt ctagagggga attgttatcc gctcacaatt
      721 cccctatagt gagtcgtatt aatttcgcgg gatcgagatc tcgatcctct acgccggacg
      781 catcgtggcc ggcatcaccg gcgccacagg tgcggttgct ggcgcctata tcgccgacat
      841 caccgatggg gaagatcggg ctcgccactt cgggctcatg agcgcttgtt tcggcgtggg
      901 tatggtggca ggccccgtgg ccgggggact gttgggcgcc atctccttgc atgcaccatt
      961 ccttgcggcg gcggtgctca acggcctcaa cctactactg ggctgcttcc taatgcagga
     1021 gtcgcataag ggagagcgtc gagatcccgg acaccatcga atggcgcaaa acctttcgcg
     1081 gtatggcatg atagcgcccg gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa
     1141 cgttatacga tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga
     1201 accaggccag ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc
     1261 tgaattacat tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg
     1321 gcgttgccac ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat
     1381 ctcgcgccga tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg
     1441 aagcctgtaa agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta
     1501 actatccgct ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg
     1561 cgttatttct tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgaag
     1621 acggtacgcg actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt
     1681 tagcgggccc attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc
     1741 tcactcgcaa tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg
     1801 gttttcaaca aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg
     1861 ccaacgatca gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg
     1921 gtgcggatat ctcggtagtg ggatacgacg ataccgaaga cagctcatgt tatatcccgc
     1981 cgttaaccac catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc
     2041 tgcaactctc tcagggccag gcggtgaagg gcaatcagct gttgcccgtc tcactggtga
     2101 aaagaaaaac caccctggcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     2161 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
     2221 attaatgtaa gttagctcac tcattaggca ccgggatctc gaccgatgcc cttgagagcc
     2281 ttcaacccag tcagctcctt ccggtgggcg cggggcatga ctatcgtcgc cgcacttatg
     2341 actgtcttct ttatcatgca actcgtagga caggtgccgg cagcgctctg ggtcattttc
     2401 ggcgaggacc gctttcgctg gagcgcgacg atgatcggcc tgtcgcttgc ggtattcgga
     2461 atcttgcacg ccctcgctca agccttcgtc actggtcccg ccaccaaacg tttcggcgag
     2521 aagcaggcca ttatcgccgg catggcggcc ccacgggtgc gcatgatcgt gctcctgtcg
     2581 ttgaggaccc ggctaggctg gcggggttgc cttactggtt agcagaatga atcaccgata
     2641 cgcgagcgaa cgtgaagcga ctgctgctgc aaaacgtctg cgacctgagc aacaacatga
     2701 atggtcttcg gtttccgtgt ttcgtaaagt ctggaaacgc ggaagtcagc gccctgcacc
     2761 attatgttcc ggatctgcat cgcaggatgc tgctggctac cctgtggaac acctacatct
     2821 gtattaacga agcgctggca ttgaccctga gtgatttttc tctggtcccg ccgcatccat
     2881 accgccagtt gtttaccctc acaacgttcc agtaaccggg catgttcatc atcagtaacc
     2941 cgtatcgtga gcatcctctc tcgtttcatc ggtatcatta cccccatgaa cagaaatccc
     3001 ccttacacgg aggcatcagt gaccaaacag gaaaaaaccg cccttaacat ggcccgcttt
     3061 atcagaagcc agacattaac gcttctggag aaactcaacg agctggacgc ggatgaacag
     3121 gcagacatct gtgaatcgct tcacgaccac gctgatgagc tttaccgcag ctgcctcgcg
     3181 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     3241 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     3301 gggtgtcggg gcgcagccat gacccagtca cgtagcgata gcggagtgta tactggctta
     3361 actatgcggc atcagagcag attgtactga gagtgcacca tatatgcggt gtgaaatacc
     3421 gcacagatgc gtaaggagaa aataccgcat caggcgctct tccgcttcct cgctcactga
     3481 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     3541 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     3601 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     3661 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     3721 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     3781 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     3841 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     3901 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     3961 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     4021 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     4081 gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     4141 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     4201 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     4261 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     4321 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     4381 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     4441 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     4501 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     4561 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     4621 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     4681 agttaatagt ttgcgcaacg ttgttgccat tgctgcaggc atcgtggtgt cacgctcgtc
     4741 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     4801 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     4861 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     4921 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     4981 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     5041 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     5101 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     5161 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     5221 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     5281 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     5341 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg aaattgtaaa
     5401 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     5461 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     5521 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     5581 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     5641 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     5701 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     5761 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     5821 gcttaatgcg ccgctacagg gcgcgtccca ttcgcca
//
