LOCUS       sequence-212334-        5910 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression for structure determination; may not be full 
            ORF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5910)
  TITLE     SGC Oxford April 2018 plasmids
REFERENCE   2  (bases 1 to 5910)
  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"
COMMENT     Sequence Label: KALRN (KALRNA-c001)
FEATURES             Location/Qualifiers
     source          1..5910
                     /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             92..121
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             146..166
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             775..792
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(845..863)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      859..906
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      943..1398
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1030..1049)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1240..1261
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1490..2305)
                     /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(1490..2305)
                     /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     2213..2232
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2297..2316
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2427..3015
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2916..2935
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3169..3186
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3201..3340
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3266..3285)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3426..3448
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3442..3633)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4442..5524)
                     /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     5486..5505
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5525..5602)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5808..5827
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gcaccatcat catcatcatc atcatcatca
      121 ttcttctggt gtagatctgg gtaccgagaa cctgtacttc caatccatgc gcaagaaaga
      181 gttcatcatg gctgaacttc tgcaaaccga gaaagcgtat gttcgcgatt tacacgaatg
      241 tctggaaacg tatctgtggg aaatgacctc aggagtggaa gagattcctc cagggattct
      301 gaacaaagag cacattatct ttggcaatat tcaggaaatc tacgactttc acaacaacat
      361 ctttctcaaa gaactcgaga aatacgaaca actgcccgaa gatgtgggtc attgctttgt
      421 gacatgggca gacaaattcc agatgtacgt cacctattgc aagaacaaac cggattcgaa
      481 tcaactgatt ctggaacatg ccggtacgtt ctttgacgaa attcagcagc gtcatggctt
      541 agccaattcc atttctagct atctgatcaa accggtacag cgtgtcacca aatatcagtt
      601 gcttctgaaa gaattgctga cttgttgcga ggaaggtaaa ggcgaattga aggatggctt
      661 agaggtgatg ctgagtgttc cgaagaaagc gaatgatgcg atgcatgtat gacagtaaag
      721 gtggatacgg atccgaattc gagctccgtc gacaagcttg cggccgcact cgagcaccac
      781 caccaccacc actgagatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct
      841 gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt
      901 tttttgctga aaggaggaac tatatccgga ttggcgaatg ggacgcgccc tgtagcggcg
      961 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     1021 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     1081 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     1141 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     1201 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     1261 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     1321 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     1381 tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     1441 tttatttttc taaatacatt caaatatgta tccgctcatg aattaattct tagaaaaact
     1501 catcgagcat caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt
     1561 gaaaaagccg tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa
     1621 gatcctggta tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc
     1681 cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg
     1741 agaatggcaa aagtttatgc atttctttcc agacttgttc aacaggccag ccattacgct
     1801 cgtcatcaaa atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga
     1861 gacgaaatac gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc
     1921 gcaggaacac tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata
     1981 cctggaatgc tgttttcccg gggatcgcag tggtgagtaa ccatgcatca tcaggagtac
     2041 ggataaaatg cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca
     2101 tctcatctgt aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg
     2161 catcgggctt cccatacaat cgatagattg tcgcacctga ttgcccgaca ttatcgcgag
     2221 cccatttata cccatataaa tcagcatcca tgttggaatt taatcgcggc ctagagcaag
     2281 acgtttcccg ttgaatatgg ctcataacac cccttgtatt actgtttatg taagcagaca
     2341 gttttattgt tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     2401 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     2461 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     2521 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
     2581 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     2641 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     2701 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     2761 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     2821 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     2881 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     2941 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     3001 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     3061 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     3121 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     3181 agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     3241 tcacaccgca atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
     3301 atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc
     3361 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac
     3421 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca
     3481 gctgcggtaa agctcatcag cgtggtcgtg aagcgattca cagatgtctg cctgttcatc
     3541 cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc
     3601 catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct
     3661 gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga
     3721 tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg tatggatgcg
     3781 gcgggaccag agaaaaatca ctcagggtca atgccagcgc ttcgttaata cagatgtagg
     3841 tgttccacag ggtagccagc agcatcctgc gatgcagatc cggaacataa tggtgcaggg
     3901 cgctgacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca ttcatgttgt
     3961 tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga
     4021 ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca acgacaggag
     4081 cacgatcatg cgcacccgtg gggccgccat gccggcgata atggcctgct tctcgccgaa
     4141 acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac
     4201 cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac
     4261 ccagagcgct gccggcacct gtcctacgag ttgcatgata aagaagacag tcataagtgc
     4321 ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa
     4381 gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg
     4441 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     4501 acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg
     4561 agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt
     4621 ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat
     4681 aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatatccgca ccaacgcgca
     4741 gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca
     4801 tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg
     4861 cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat
     4921 gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga
     4981 tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg
     5041 agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat
     5101 tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca
     5161 gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc
     5221 ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg
     5281 ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca
     5341 acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca
     5401 tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc
     5461 gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt
     5521 tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg
     5581 ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga ctcctgcatt
     5641 aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca
     5701 tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg
     5761 aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg
     5821 atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg
     5881 tagaggatcg agatctcgat cccgcgaaat
//
