LOCUS       sequence-181159-        5967 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 5967)
  AUTHORS   Schwartz TU, Schmidt D, Brohawn SG, Blobel G
  TITLE     Homodimerization of the G protein SRbeta in the nucleotide-free 
            state involves proline cis/trans isomerization in the switch II 
            region.
  JOURNAL   Proc Natl Acad Sci U S A. 2006 May 2. 103(18):6823-8.
  PUBMED    16627619
REFERENCE   2  (bases 1 to 5967)
  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: "Proc Natl 
            Acad Sci U S A. 2006 May 2. 103(18):6823-8."
COMMENT     Sequence Label: pHsSR beta deltaN Flag
FEATURES             Location/Qualifiers
     source          1..5967
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      3..50
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      87..542
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(174..193)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     384..405
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(634..1449)
                     /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(634..1449)
                     /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     1357..1376
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1441..1460
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1571..2159
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2060..2079
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2313..2330
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2345..2484
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2410..2429)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2570..2592
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2586..2777)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3586..4668)
                     /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     4630..4649
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4669..4746)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     4952..4971
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5055..5074
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5055..5073
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5074..5098
                     /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             5113..5135
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5154..5171
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5181..5198
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             5850..5873
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             5886..5903
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(join(5956..5967,1..7))
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
ORIGIN
        1 aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag
       61 gaactatatc cggattggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg
      121 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
      181 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
      241 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
      301 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
      361 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
      421 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
      481 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat
      541 ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
      601 cattcaaata tgtatccgct catgaattaa ttcttagaaa aactcatcga gcatcaaatg
      661 aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa gccgtttctg
      721 taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct ggtatcggtc
      781 tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt caaaaataag
      841 gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg gcaaaagttt
      901 atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat caaaatcact
      961 cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgagacgaa atacgcgatc
     1021 gctgttaaaa ggacaattac aaacaggaat cgaatgcaac cggcgcagga acactgccag
     1081 cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga atgctgtttt
     1141 cccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa aatgcttgat
     1201 ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat ctgtaacatc
     1261 attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg gcttcccata
     1321 caatcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt tatacccata
     1381 taaatcagca tccatgttgg aatttaatcg cggcctagag caagacgttt cccgttgaat
     1441 atggctcata acaccccttg tattactgtt tatgtaagca gacagtttta ttgttcatga
     1501 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     1561 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     1621 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     1681 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     1741 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     1801 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     1861 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     1921 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     1981 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     2041 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     2101 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     2161 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     2221 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     2281 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     2341 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcaatggtg
     2401 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg
     2461 ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga
     2521 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     2581 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca
     2641 tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg
     2701 agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt
     2761 ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg
     2821 ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga acatgcccgg
     2881 ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga ccagagaaaa
     2941 atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc acagggtagc
     3001 cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga cttccgcgtt
     3061 tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca ggtcgcagac
     3121 gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca
     3181 gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat catgcgcacc
     3241 cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt ggtggcggga
     3301 ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag cgacaggccg
     3361 atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc
     3421 acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac gatagtcatg
     3481 ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat
     3541 cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc
     3601 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     3661 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
     3721 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
     3781 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
     3841 ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat
     3901 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
     3961 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
     4021 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
     4081 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
     4141 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
     4201 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
     4261 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
     4321 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
     4381 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
     4441 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
     4501 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
     4561 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
     4621 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
     4681 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
     4741 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
     4801 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc
     4861 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
     4921 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
     4981 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct
     5041 cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata acaattcccc
     5101 tctagaaata attttgttta actttaagaa ggagatatac catgggcagc agccatcatc
     5161 atcatcatca cagcagcggc ctggtgccgc gcggcagcca tatgagcaga aggagcagtc
     5221 agagagctgt tcttcttgtt ggcctttgtg attccgggaa aacgttgctc tttgtcaggt
     5281 tgttaacagg cctttataga gacactcaga cgtccattac tgacagctgt gctgtataca
     5341 gagtcaacaa taacaggggc aatagtctga ccttgattga ccttcccggc catgagagtt
     5401 tgaggcttca gttcttagag cggtttaagt cttcagccag ggctattgtg tttgttgtgg
     5461 atagtgcagc attccagcga gaggtgaaag atgtggctga gtttctgtat caagtcctca
     5521 ttgacagtat gggtctgaag aatacaccat cattcttaat agcctgcaat aagcaagata
     5581 ttgcaatggc aaaatcagca aagttaattc aacagcagct ggagaaagaa ctcaacacct
     5641 tacgagttac ccgttctgct gcccccagca cactggacag ttccagcact gcccctgctc
     5701 agctggggaa gaaaggcaaa gagtttgaat tctcacagtt gcccctcaaa gtggagttcc
     5761 tggagtgcag tgccaagggt ggaagagggg acgtgggctc tgctgacatc caggacttgg
     5821 agaaatggct ggctaaaatt gccaagcttg attacaagga tgacgacgat aagtgataac
     5881 tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga aaggaagctg
     5941 agttggctgc tgccaccgct gagcaat
//
