LOCUS       Exported                5917 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Bacterial expression for structure determination; may not be full 
            ORF.
ACCESSION   .
VERSION     .
KEYWORDS    SHC1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5917)
  TITLE     SGC Oxford
REFERENCE   2  (bases 1 to 5917)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5917
                     /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             778..795
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(848..866)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      862..909
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      946..1401
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1033..1052)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1243..1264
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1494..2309)
                     /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     2217..2236
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2301..2320
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2431..3019
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2920..2939
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3173..3190
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3205..3347
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3273..3292)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3433..3455
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3449..3640)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4449..5531)
                     /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     5493..5512
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5532..5609)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5815..5834
                     /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 atccatgggc cagcttgggg gcgaggagtg
      181 gacccgccac gggagctttg tcaataagcc cacgcggggc tggctgcatc ccaacgacaa
      241 agtcatggga cccggggttt cctacttggt tcggtacatg ggttgtgtgg aggtcctcca
      301 gtcaatgcgt gccctggact tcaacacccg gactcaggtc accagggagg ccatcagtct
      361 ggtgtgtgag gctgtgccgg gtgctaaggg ggcgacaagg aggagaaagc cctgtagccg
      421 cccgctcagc tctatcctgg ggaggagtaa cctgaaattt gctggaatgc caatcactct
      481 caccgtctcc accagcagcc tcaacctcat ggccgcagac tgcaaacaga tcatcgccaa
      541 ccaccacatg caatctatct catttgcatc cggcggggat ccggacacag ccgagtatgt
      601 cgcctatgtt gccaaagacc ctgtgaatca gagagcctgc cacattctgg agtgtcccga
      661 agggcttgcc caggatgtca tcagcaccat tggccaggcc ttcgagttgc gctgacagta
      721 aaggtggata cggatccgaa ttcgagctcc gtcgacaagc ttgcggccgc actcgagcac
      781 caccaccacc accactgaga tccggctgct aacaaagccc gaaaggaagc tgagttggct
      841 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
      901 ggttttttgc tgaaaggagg aactatatcc ggattggcga atgggacgcg ccctgtagcg
      961 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     1021 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     1081 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     1141 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
     1201 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     1261 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     1321 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     1381 aaatattaac gtttacaatt tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     1441 tttgtttatt tttctaaata cattcaaata tgtatccgct catgaattaa ttcttagaaa
     1501 aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat
     1561 ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg
     1621 gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat
     1681 ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc
     1741 ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta
     1801 cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga
     1861 gcgagacgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac
     1921 cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct
     1981 aatacctgga atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga
     2041 gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg
     2101 accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct
     2161 ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg
     2221 cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag
     2281 caagacgttt cccgttgaat atggctcata acaccccttg tattactgtt tatgtaagca
     2341 gacagtttta ttgttcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     2401 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     2461 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     2521 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct
     2581 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     2641 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     2701 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     2761 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     2821 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     2881 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     2941 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     3001 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     3061 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     3121 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     3181 agtgagcgag gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg
     3241 tatttcacac cgcatatatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3301 agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc
     3361 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     3421 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     3481 cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct
     3541 gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa
     3601 agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg
     3661 gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg
     3721 ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat
     3781 ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag
     3841 atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg
     3901 tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc
     3961 atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta
     4021 tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg
     4081 acaggagcac gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg gcctgcttct
     4141 cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc
     4201 cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga
     4261 aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca
     4321 taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg
     4381 ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg
     4441 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4501 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc
     4561 accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc
     4621 aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc
     4681 gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca
     4741 acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca
     4801 accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg
     4861 gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga
     4921 tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac
     4981 agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct
     5041 tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc
     5101 ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta
     5161 atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg
     5221 acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat
     5281 ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca
     5341 atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc
     5401 tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc
     5461 accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt
     5521 actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg
     5581 cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc
     5641 ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa
     5701 tggtgcatgc aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc
     5761 cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc cgatcttccc catcggtgat
     5821 gtcggcgata taggcgccag caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg
     5881 tccggcgtag aggatcgaga tctcgatccc gcgaaat
//
