LOCUS       sequence-372319-        4682 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of heat-resistant obscure (Hero) protein Hero9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4682)
  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 4682)
  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-Hero9
FEATURES             Location/Qualifiers
     source          1..4682
                     /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             522..545
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             546..563
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             566..577
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     3'UTR           645..789
                     /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(830..852)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     952..971
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(988..1443)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1125..1146)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1337..1356
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1543..1647
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1648..1716,1717..2508)
                     /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(1866..1885)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2679..3267
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3168..3187
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3439..4521)
                     /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     4483..4502
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4522..4599)
                     /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 tgatgtcggg ccccaacgga gacctgggga
      301 tgccggtgga ggcgggagcg gaaggcgagg aggacggctt cggggaagca gaatacgctg
      361 ccatcaactc catgctggac cagatcaact cctgtctgga ccacctggag gagaagaatg
      421 accacctcca cgcccgcctc caggagctgc tggagtccaa ccggcagaca cgcctggagt
      481 tccagcagca gctcggggag gcccccagtg atgccagccc cgactacaaa gacgatgacg
      541 acaagcatca tcatcatcat cattaatcga aggtaggcat atggagctcg gtaccctcga
      601 gggatccgaa ttcaagcttg tcgacctgca gtctagatag gtaatctctg cttaaaagca
      661 cagaatctaa gatccctgcc atttggcggg gattttttta tttgttttca ggaaataaat
      721 aatcgatcgc gtaataaaat ctattattat ttttgtgaag aataaatttg ggtgcaatga
      781 gaatgcgcag gccctttcgt ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca
      841 tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc
      901 gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat gcggcatcag
      961 agcagattgt actgagagtg caccataaaa ttgtaaacgt taatattttg ttaaaattcg
     1021 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     1081 cttataaatc aaaagaatag cccgagatag ggttgagtgt tgttccagtt tggaacaaga
     1141 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     1201 atggcccact acgtgaacca tcacccaaat caagtttttt ggggtcgagg tgccgtaaag
     1261 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     1321 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     1381 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     1441 cgtactatgg ttgctttgac gtatgcggtg tgaaataccg cacagatgcg taaggagaaa
     1501 ataccgcatc aggcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     1561 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     1621 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     1681 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     1741 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     1801 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     1861 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     1921 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     1981 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     2041 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     2101 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     2161 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     2221 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     2281 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     2341 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     2401 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     2461 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     2521 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     2581 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     2641 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     2701 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     2761 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     2821 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     2881 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     2941 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     3001 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     3061 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     3121 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     3181 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     3241 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     3301 gccttttgct ggccttttgc tcacatagtc atgccccgcg cccaccggaa ggagctgact
     3361 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
     3421 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3481 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
     3541 ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
     3601 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
     3661 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
     3721 atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
     3781 atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg
     3841 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
     3901 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
     3961 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
     4021 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     4081 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
     4141 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     4201 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     4261 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     4321 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     4381 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     4441 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     4501 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
     4561 tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct
     4621 tatgcgactc ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg
     4681 cc
//
