LOCUS       sequence-372316-        4766 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of heat-resistant obscure (Hero) protein 
            Hero13.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4766)
  AUTHORS   Tsuboyama K, Osaki T, Matsuura-Suzuki E, Kozuka-Hata H, Okada Y, 
            Oyama M, Ikeuchi Y, Iwasaki S, Tomari Y
  TITLE     A widespread family of heat-resistant obscure (Hero) proteins 
            protect against protein instability and aggregation.
  JOURNAL   PLoS Biol. 2020 Mar 12;18(3):e3000632. doi: 
            10.1371/journal.pbio.3000632. eCollection 2020 Mar.
  PUBMED    32163402
REFERENCE   2  (bases 1 to 4766)
  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: "PLoS 
            Biol."; date: "2020-03-12"; volume: "18(3):e3000632. doi"; pages: " 
            10.1371/journal.pbio.3000632. eCollection 2020 Mar"
COMMENT     Sequence Label: pCold I-Hero13
FEATURES             Location/Qualifiers
     source          1..4766
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        17..83
                     /gene="cspA"
                     /label=cspA promoter
                     /note="promoter of the E. coli cold shock protein cspA gene
                     (Mitta et al., 1997)"
     protein_bind    86..102
                     /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)."
     5'UTR           124..257
                     /gene="cspA"
                     /label=cspA 5'UTR
                     /note="5'UTR of the E. coli cold shock protein cspA gene 
                     (Mitta et al., 1997)"
     CDS             258..272
                     /codon_start=1
                     /gene="cspA"
                     /product="translation enhancing element for E. coli (Qing 
                     et al., 2004)"
                     /label=TEE
                     /translation="MNHKV"
     CDS             606..629
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             630..647
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             650..661
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     3'UTR           729..873
                     /gene="cspA"
                     /label=cspA 3'UTR
                     /note="3'UTR of the E. coli cold shock protein cspA gene 
                     (Mitta et al., 1997)"
     primer_bind     complement(914..936)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1036..1055
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(1072..1527)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1209..1230)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1421..1440
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1627..1731
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1732..1800,1801..2592)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1950..1969)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2763..3351
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3252..3271
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3523..4605)
                     /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     4567..4586
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4606..4683)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 gcaaggaatg gtgtggccga ttaatcataa atatgaaaaa taattgttgc atcacccgcc
       61 aatgcgtggc ttaatgcaca tcaaattgtg agcggataac aatttgatgt gctagcgcat
      121 atccagtgta gtaaggcaag tcccttcaag agttatcgtt gatacccctc gtagtgcaca
      181 ttcctttaac gcttcaaaat ctgtaaagca cgccatatcg ccgaaaggca cacttaatta
      241 ttaagaggta atacaccatg aatcacaaag tgatggcgaa catccaccag gaaaacgaag
      301 agatggaaca gcccatgcag aatggcgaag aagatcgtcc tcttggcgga ggtgaaggcc
      361 atcaaccagc aggcaatcgt cgtgggcaag ctcgccgttt agcgccgaac tttcgctggg
      421 ccattccgaa tcgccagatc aacgatggta tgggtggtga tggggatgac atggagatct
      481 tcatggaaga gatgcgggaa attcgccgta aactgcgcga actccagttg cgcaattgtc
      541 tgcgcattct gatgggcgaa ctgagcaacc accatgacca tcacgacgag ttttgcctga
      601 tgccggacta caaagacgat gacgacaagc atcatcatca tcatcattaa tcgaaggtag
      661 gcatatggag ctcggtaccc tcgagggatc cgaattcaag cttgtcgacc tgcagtctag
      721 ataggtaatc tctgcttaaa agcacagaat ctaagatccc tgccatttgg cggggatttt
      781 tttatttgtt ttcaggaaat aaataatcga tcgcgtaata aaatctatta ttatttttgt
      841 gaagaataaa tttgggtgca atgagaatgc gcaggccctt tcgtctcgcg cgtttcggtg
      901 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
      961 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     1021 gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat aaaattgtaa
     1081 acgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     1141 aataggccga aatcggcaaa atcccttata aatcaaaaga atagcccgag atagggttga
     1201 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     1261 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc aaatcaagtt
     1321 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     1381 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     1441 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     1501 cgcttaatgc gccgctacag ggcgcgtact atggttgctt tgacgtatgc ggtgtgaaat
     1561 accgcacaga tgcgtaagga gaaaataccg catcaggcgt caggtggcac ttttcgggga
     1621 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     1681 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     1741 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     1801 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     1861 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     1921 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     1981 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     2041 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     2101 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     2161 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     2221 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     2281 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     2341 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     2401 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     2461 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     2521 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     2581 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     2641 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     2701 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     2761 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     2821 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     2881 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     2941 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     3001 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     3061 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     3121 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     3181 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     3241 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     3301 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     3361 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat agtcatgccc
     3421 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     3481 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     3541 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     3601 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     3661 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     3721 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     3781 atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact cggtaatggc
     3841 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     3901 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     3961 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     4021 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
     4081 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
     4141 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
     4201 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
     4261 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
     4321 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
     4381 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
     4441 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
     4501 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
     4561 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
     4621 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     4681 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag cccagtagta
     4741 ggttgaggcc gttgagcacc gccgcc
//
