LOCUS       sequence-360467-        4388 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of human growth hormone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4388)
  AUTHORS   Eastwood TA, Baker K, Streather BR, Allen N, Wang L, Botchway SW, 
            Brown IR, Hiscock JR, Lennon C, Mulvihill DP
  TITLE     High-yield vesicle-packaged recombinant protein production from E. 
            coli.
  JOURNAL   Cell Rep Methods. 2023 Feb 2;3(2):100396. doi: 
            10.1016/j.crmeth.2023.100396. eCollection 2023 Feb 27.
  PUBMED    36936078
REFERENCE   2  (bases 1 to 4388)
  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: "Cell Rep 
            Methods."; date: "2023-02-2"; volume: "3(2):100396. doi"; pages: " 
            10.1016/j.crmeth.2023.100396. eCollection 2023 Feb 27"
COMMENT     Sequence Label: pRSFDuet-1_hGH
FEATURES             Location/Qualifiers
     source          1..4388
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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)."
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        214..232
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    233..257
                     /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)."
     CDS             892..909
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             925..969
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1007..1025)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1021..1068
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1298..2113)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1298..2113)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2021..2040
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2114..2205)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2221..2970)
                     /direction=LEFT
                     /label=RSF ori
                     /note="Plasmids containing the RSF 1030 origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     primer_bind     complement(2231..2248)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2471..2490)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3166..4248)
                     /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     4210..4229
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4249..4326)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(4372..4388,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cgaattcgag
      121 ctcggcgcgc ctgcaggtcg acaagcttgc ggccgcataa tgcttaagtc gaacagaaag
      181 taatcgtatt gtacacggcc gcataatcga aattaatacg actcactata ggggaattgt
      241 gagcggataa caattcccca tcttagtata ttagttaagt ataagaagga gatatacata
      301 catatgttcc cgacgattcc gctgtctcgt ctgttcgata acgctatgct gcgtgcacat
      361 cgtctgcacc agctggcatt cgacacttat caagaattcg aggaggccta catcccgaaa
      421 gaacagaaat acagcttcct gcagaacccg caaacctccc tgtgcttcag cgaatccatc
      481 ccgactccgt ctaaccgcga agaaacccag cagaaatcta atctggaact gctgcgtatt
      541 agcctgctgc tgatccagtc ttggctggaa ccggtgcaat tcctgcgcag cgtcttcgcg
      601 aactccctgg tatatggcgc ttccgactct aacgtatacg atctgctgaa agacctggag
      661 gagggtatcc agaccctgat gggccgcctg gaggacggca gcccgcgtac tggccagatt
      721 tttaaacaga cttactctaa attcgacacc aactcccata atgatgacgc cctgctgaaa
      781 aactatggtc tgctgtactg cttccgtaaa gacatggaca aagttgagac cttcctgcgt
      841 attgttcagt gtcgctctgt tgagggttct tgcggcttct aatgaggtac ccatcatcat
      901 catcatcatt aactcgagtc tggtaaagaa accgctgctg cgaaatttga acgccagcac
      961 atggactcgt ctactagcgc agcttaatta acctaggctg ctgccaccgc tgagcaataa
     1021 ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaacctcag
     1081 gcatttgaga agcacacggt cacactgctt ccggtagtca ataaaccggt aaaccagcaa
     1141 tagacataag cggctattta acgaccctgc cctgaaccga cgacaagctg acgaccgggt
     1201 ctccgcaagt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     1261 aatacattca aatatgtatc cgctcatgaa ttaattctta gaaaaactca tcgagcatca
     1321 aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt
     1381 tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga tcctggtatc
     1441 ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa
     1501 taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaaa
     1561 gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg tcatcaaaat
     1621 cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc
     1681 ggtcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc aggaacactg
     1741 ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc tggaatgctg
     1801 ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg ataaaatgct
     1861 tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc tcatctgtaa
     1921 catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc
     1981 catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc catttatacc
     2041 catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac gtttcccgtt
     2101 gaatatggct catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     2161 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggc atgcagcgct
     2221 cttccgcttc ctcgctcact gactcgctac gctcggtcgt tcgactgcgg cgagcggtgt
     2281 cagctcactc aaaagcggta atacggttat ccacagaatc aggggataaa gccggaaaga
     2341 acatgtgagc aaaaagcaaa gcaccggaag aagccaacgc cgcaggcgtt tttccatagg
     2401 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gccagaggtg gcgaaacccg
     2461 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     2521 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     2581 tctcatagct cacgctgttg gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2641 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     2701 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccattgg taactgattt
     2761 agaggacttt gtcttgaagt tatgcacctg ttaaggctaa actgaaagaa cagattttgg
     2821 tgagtgcggt cctccaaccc acttaccttg gttcaaagag ttggtagctc agcgaacctt
     2881 gagaaaacca ccgttggtag cggtggtttt tctttattta tgagatgatg aatcaatcgg
     2941 tctatcaagt caacgaacag ctattccgtt actctagatt tcagtgcaat ttatctcttc
     3001 aaatgtagca cctgaagtca gccccatacg atataagttg taattctcat gttagtcatg
     3061 ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat
     3121 cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc
     3181 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     3241 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
     3301 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
     3361 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
     3421 ggtatcgtcg tatcccacta ccgagatgtc cgcaccaacg cgcagcccgg actcggtaat
     3481 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
     3541 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
     3601 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
     3661 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
     3721 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
     3781 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
     3841 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
     3901 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
     3961 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
     4021 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
     4081 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
     4141 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
     4201 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
     4261 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
     4321 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaaa ttaatacgac
     4381 tcactata
//
