LOCUS       sequence-313989-        5786 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 5786)
  AUTHORS   Chivers CE, Crozat E, Chu C, Moy VT, Sherratt DJ, Howarth M
  TITLE     A streptavidin variant with slower biotin dissociation and increased
            mechanostability.
  JOURNAL   Nat Methods. 2010 May;7(5):391-3. doi: 10.1038/nmeth.1450. Epub 2010
            Apr 11.
  PUBMED    20383133
REFERENCE   2  (bases 1 to 5786)
  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"; doi: 
            "10.1038/nmeth.1450"; journalName: "Nat Methods"; date: "2010-05"; 
            volume: "7"; issue: "5"; pages: "391-3"
COMMENT     Sequence Label: pET21a-Traptavidin
FEATURES             Location/Qualifiers
     source          1..5786
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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)."
     RBS             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             473..490
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             518..535
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(588..606)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      602..649
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      686..1141
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(773..792)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     983..1004
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1167..1271
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1272..1340,1341..2132)
                     /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     complement(1490..1509)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2303..2891
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2792..2811
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3045..3062
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3077..3216
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3142..3161)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3302..3324
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3318..3509)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4318..5400)
                     /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     5362..5381
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5401..5478)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5684..5703
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat ggctgaagct ggtatcaccg gcacctggta
      121 caaccagctg ggatccacct tcatcgttac cgctggtgct gacggtgctc tgaccggtac
      181 ctacgaatcc gctgttggta acgctgaagg cgattacgtt ctgaccggtc gttacgactc
      241 cgctccggct accgacggtt ccggaaccgc tctgggttgg accgttgctt ggaaaaacaa
      301 ctaccgtaac gctcactccg ctaccacctg gtctggccag tacgttggtg gtgctgaagc
      361 tcgtatcaac acccagtggt tgttgacctc cggcaccacc gaagccaacg cgtggaaatc
      421 caccctggtt ggtcacgaca ccttcaccaa agttaaaccg tccgctgctt cccatcacca
      481 tcaccaccat taataaaagc ttgcggccgc actcgagcac caccaccacc accactgaga
      541 tccggctgct aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata
      601 actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg
      661 aactatatcc ggattggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt
      721 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
      781 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
      841 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
      901 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
      961 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     1021 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     1081 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt
     1141 taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     1201 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     1261 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     1321 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     1381 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     1441 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     1501 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     1561 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     1621 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     1681 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     1741 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     1801 acaccacgat gcctgcagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     1861 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     1921 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     1981 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     2041 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     2101 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     2161 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     2221 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     2281 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     2341 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     2401 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     2461 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     2521 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     2581 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     2641 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     2701 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     2761 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     2821 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     2881 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     2941 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     3001 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     3061 aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3121 accgcaatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac
     3181 actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct
     3241 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     3301 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg
     3361 cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg
     3421 tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg
     3481 ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc
     3541 atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat
     3601 gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg
     3661 gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt
     3721 ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct
     3781 gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct
     3841 caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca
     3901 ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg
     3961 atcatgcgca cccgtggggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt
     4021 ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca
     4081 agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag
     4141 agcgctgccg gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg
     4201 acgatagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc
     4261 atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc gttgcgctca
     4321 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     4381 gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca ccagtgagac
     4441 gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac
     4501 gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg ggatataaca
     4561 tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa cgcgcagccc
     4621 ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc
     4681 agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg acatggcact
     4741 ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat atttatgcca
     4801 gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca gcgcgatttg
     4861 ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt catgggagaa
     4921 aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg gaacattagt
     4981 gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa tgatcagccc
     5041 actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga cgccgcttcg
     5101 ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc
     5161 gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga
     5221 ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc
     5281 cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga
     5341 aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac
     5401 attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt
     5461 gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga
     5521 agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca
     5581 aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac
     5641 aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat
     5701 aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga
     5761 ggatcgagat ctcgatcccg cgaaat
//
