LOCUS       sequence-388009-        5410 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of HMfA K31A E35A in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5410)
  AUTHORS   Erkelens AM, Henneman B, van der Valk RA, Kirolos NCS, Dame RT
  TITLE     Specific DNA binding of archaeal histones HMfA and HMfB.
  JOURNAL   Front Microbiol. 2023 Apr 18;14:1166608. doi: 
            10.3389/fmicb.2023.1166608. eCollection 2023.
  PUBMED    37143534
REFERENCE   2  (bases 1 to 5410)
  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: "Front 
            Microbiol."; date: "2023-04-18"; volume: "14:1166608. doi"; pages: "
            10.3389/fmicb.2023.1166608. eCollection 2023"
COMMENT     Sequence Label: pRD323
FEATURES             Location/Qualifiers
     source          1..5410
                     /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"
     RBS             347..369
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    384..408
                     /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(408..427)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(409..427)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(515..534)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        740..817
                     /gene="lacI"
                     /label=lacI promoter
     CDS             818..1900
                     /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(837..856)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             2709..2900
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2894..2916)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3002..3141
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3057..3076
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3156..3173)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3327..3915)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3407..3426)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4026..4045)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4037..4852
                     /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             4037..4852
                     /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     complement(4110..4129)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(4944..5399)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5081..5102)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5293..5312
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagt tatttgaaca tttttcgagc taattcaatg tcttctgcct ttattgtttt
      181 tcttcccgca tgttttgcta atttcacagc ttctgaagct atttcttcgc ccatcgcttc
      241 taaaaccgct gctaatgcaa ttcttgcatc gtcactaact ctttcagcac cagcgttttt
      301 tatgattctt ccgattgggg caattggtaa ctcacccata tgtatatctc cttcttaaag
      361 ttaaacaaaa ttatttctag aggggaattg ttatccgctc acaattcccc tatagtgagt
      421 cgtattaatt tcgcgggatc gagatcgatc tcgatcctct acgccggacg catcgtggcc
      481 ggcatcaccg gcgccacagg tgcggttgct ggcgcctata tcgccgacat caccgatggg
      541 gaagatcggg ctcgccactt cgggctcatg agcgcttgtt tcggcgtggg tatggtggca
      601 ggccccgtgg ccgggggact gttgggcgcc atctccttgc atgcaccatt ccttgcggcg
      661 gcggtgctca acggcctcaa cctactactg ggctgcttcc taatgcagga gtcgcataag
      721 ggagagcgtc gagatcccgg acaccatcga atggcgcaaa acctttcgcg gtatggcatg
      781 atagcgcccg gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa cgttatacga
      841 tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga accaggccag
      901 ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc tgaattacat
      961 tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg gcgttgccac
     1021 ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga
     1081 tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa
     1141 agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta actatccgct
     1201 ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg cgttatttct
     1261 tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgaag acggtacgcg
     1321 actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc
     1381 attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc tcactcgcaa
     1441 tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg gttttcaaca
     1501 aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg ccaacgatca
     1561 gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg gtgcggacat
     1621 ctcggtagtg ggatacgacg ataccgaaga cagctcatgt tatatcccgc cgttaaccac
     1681 catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc tgcaactctc
     1741 tcagggccag gcggtgaagg gcaatcagct gttgcccgtc tcactggtga aaagaaaaac
     1801 caccctggcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     1861 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtaa
     1921 gttagctcac tcattaggca ccgggatctc gaccgatgcc cttgagagcc ttcaacccag
     1981 tcagctcctt ccggtgggcg cggggcatga ctatcgtcgc cgcacttatg actgtcttct
     2041 ttatcatgca actcgtagga caggtgccgg cagcgctctg ggtcattttc ggcgaggacc
     2101 gctttcgctg gagcgcgacg atgatcggcc tgtcgcttgc ggtattcgga atcttgcacg
     2161 ccctcgctca agccttcgtc actggtcccg ccaccaaacg tttcggcgag aagcaggcca
     2221 ttatcgccgg catggcggcc ccacgggtgc gcatgatcgt gctcctgtcg ttgaggaccc
     2281 ggctaggctg gcggggttgc cttactggtt agcagaatga atcaccgata cgcgagcgaa
     2341 cgtgaagcga ctgctgctgc aaaacgtctg cgacctgagc aacaacatga atggtcttcg
     2401 gtttccgtgt ttcgtaaagt ctggaaacgc ggaagtcagc gccctgcacc attatgttcc
     2461 ggatctgcat cgcaggatgc tgctggctac cctgtggaac acctacatct gtattaacga
     2521 agcgctggca ttgaccctga gtgatttttc tctggtcccg ccgcatccat accgccagtt
     2581 gtttaccctc acaacgttcc agtaaccggg catgttcatc atcagtaacc cgtatcgtga
     2641 gcatcctctc tcgtttcatc ggtatcatta cccccatgaa cagaaatccc ccttacacgg
     2701 aggcatcagt gaccaaacag gaaaaaaccg cccttaacat ggcccgcttt atcagaagcc
     2761 agacattaac gcttctggag aaactcaacg agctggacgc ggatgaacag gcagacatct
     2821 gtgaatcgct tcacgaccac gctgatgagc tttaccgcag ctgcctcgcg cgtttcggtg
     2881 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     2941 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     3001 gcgcagccat gacccagtca cgtagcgata gcggagtgta tactggctta actatgcggc
     3061 atcagagcag attgtactga gagtgcacca ttgcggtgtg aaataccgca cagatgcgta
     3121 aggagaaaat accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     3181 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     3241 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     3301 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     3361 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     3421 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     3481 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     3541 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     3601 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     3661 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     3721 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt
     3781 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     3841 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     3901 aaaaaaggat ttcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     3961 gaaaactcac gttaagggat tttggtcatg aacaataaaa ctgtctgctt acataaacag
     4021 taatacaagg ggtgttatga gccatattca acgggaaacg tcttgctcta ggccgcgatt
     4081 aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca
     4141 atcaggtgcg acaatctatc gattgtatgg gaagcccgat gcgccagagt tgtttctgaa
     4201 acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct
     4261 gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg
     4321 gttactcacc actgcgatcc ccgggaaaac agcattccag gtattagaag aatatcctga
     4381 ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc
     4441 tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg
     4501 aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt
     4561 tgaacaagtc tggaaagaaa tgcataaact tttgccattc tcaccggatt cagtcgtcac
     4621 tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat
     4681 tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg
     4741 cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa
     4801 tcctgatatg aataaattgc agtttcattt gatgctcgat gagtttttct aagaattaat
     4861 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     4921 catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa aattcgcgtt
     4981 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     5041 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     5101 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     5161 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     5221 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     5281 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     5341 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     5401 ccattcgcca
//
