LOCUS       Exported                6258 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Bacterial expression for structure determination; may not be full 
            ORF.
ACCESSION   .
VERSION     .
KEYWORDS    STK10 (STK10A-c043)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6258)
  TITLE     SGC Oxford April 2018 plasmids
REFERENCE   2  (bases 1 to 6258)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6258
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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..109
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             134..154
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1123..1140
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1193..1211)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1207..1254
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1291..1746
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1378..1397)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1588..1609
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1838..2653)
                     /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     2561..2580
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2645..2664
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2775..3363
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3264..3283
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3517..3534
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3549..3688
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3614..3633)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3774..3796
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3790..3981)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4790..5872)
                     /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     5834..5853
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5873..5950)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6156..6175
                     /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 catcatcatt cttctggtgt
      121 agatctgggt accgagaacc tgtacttcca atccatgaga aagtcccgcg aatatgagca
      181 cgtccgccgc gacctggacc ccaacgaggt gtgggagatc gtgggcgagc tgggcgacgg
      241 cgccttcggc aaggtttaca aggccaagaa taaggagacg ggtgctttgg ctgcggccaa
      301 agtcattgaa accaagagtg aggaggagct ggaggactac atcgtggaga ttgagatcct
      361 ggccacctgc gaccacccct acattgtgaa gctcctggga gcctactatc acgacgggaa
      421 gctgtggatc atgattgagt tctgtccagg gggagccgtg gacgccatca tgctggagct
      481 ggacagaggc ctcacggagc cccagataca ggtggtttgc cgccagatgc tagaagccct
      541 caacttcctg cacagcaaga ggatcatcca ccgagatctg aaagctggca acgtgctgat
      601 gaccctcgag ggagacatca ggctggctga ctttggtgtg tctgccaaga atctgaagac
      661 tctacagaaa cgagattcct tcatcggcac gccttactgg atggcccccg aggtggtcat
      721 gtgtgagacc atgaaagaca cgccctacga ctacaaagcc gacatctggt ccctgggcat
      781 cacgctgatt gagatggccc agatcgagcc gccacaccac gagctcaacc ccatgcgggt
      841 cctgctagcg atcgccgcgt cagaccctcc cacgctgctc acgccctcca agtggtctgt
      901 agagttccgt gacttcctgg cgatagccct ggataagaac ccagaaaccc gacccagtgc
      961 cgcgcagctg ctggagcatc ccttcgtcag cagcatcacc agtaacaagg ctctgcggga
     1021 gctggtggct gaggccaagg ccgaggtgat ggaagagtga cagtaaaggt ggatacggat
     1081 ccgaattcga gctccgtcga caagcttgcg gccgcactcg agcaccacca ccaccaccac
     1141 tgagatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag
     1201 caataactag cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa
     1261 ggaggaacta tatccggatt ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
     1321 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     1381 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     1441 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     1501 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     1561 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     1621 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     1681 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     1741 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     1801 aatacattca aatatgtatc cgctcatgaa ttaattctta gaaaaactca tcgagcatca
     1861 aatgaaactg caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt
     1921 tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga tcctggtatc
     1981 ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa
     2041 taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaaa
     2101 gtttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg tcatcaaaat
     2161 cactcgcatc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc
     2221 gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc aggaacactg
     2281 ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc tggaatgctg
     2341 ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg ataaaatgct
     2401 tgatggtcgg aagaggcata aattccgtca gccagtttag tctgaccatc tcatctgtaa
     2461 catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc
     2521 catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc catttatacc
     2581 catataaatc agcatccatg ttggaattta atcgcggcct agagcaagac gtttcccgtt
     2641 gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt tttattgttc
     2701 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     2761 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     2821 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     2881 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     2941 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     3001 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     3061 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     3121 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     3181 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     3241 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     3301 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     3361 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     3421 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     3481 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     3541 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat
     3601 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct
     3661 atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc
     3721 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     3781 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag
     3841 ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc
     3901 gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc
     3961 ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt
     4021 aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc
     4081 ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag
     4141 aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg
     4201 tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg
     4261 cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc
     4321 agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta
     4381 accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg
     4441 cacccgtggg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc
     4501 gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag
     4561 gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc
     4621 cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt
     4681 catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg
     4741 agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc
     4801 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4861 aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca
     4921 gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt
     4981 gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt
     5041 cttcggtatc gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg
     5101 taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa
     5161 cgatgccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc
     5221 cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca
     5281 gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac
     5341 ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac
     5401 tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag
     5461 cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc
     5521 gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca
     5581 tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt
     5641 gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc
     5701 ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca
     5761 ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct
     5821 gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca
     5881 ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt
     5941 cgatggtgtc cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc
     6001 agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg
     6061 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc
     6121 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca
     6181 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag
     6241 atctcgatcc cgcgaaat
//
