LOCUS       Exported                5934 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Bacterial expression for structure determination; may not be full 
            ORF.
ACCESSION   .
VERSION     .
KEYWORDS    TRIM33 (TRIM33B-c100)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5934)
  TITLE     SGC Oxford 2017
REFERENCE   2  (bases 1 to 5934)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5934
                     /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             799..816
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(869..887)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      883..930
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      967..1422
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1054..1073)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1264..1285
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1514..2329)
                     /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     2237..2256
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2321..2340
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2451..3039
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2940..2959
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3193..3210
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3225..3364
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3290..3309)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3450..3472
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3466..3657)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4466..5548)
                     /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     5510..5529
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5549..5626)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5832..5851
                     /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 atccatggat gatgacccaa atgaagactg
      181 gtgtgctgtc tgccaaaacg gaggagatct cttgtgctgc gaaaaatgtc caaaggtctt
      241 tcatctaact tgtcatgttc caacactact tagctttcca agtggggact ggatatgcac
      301 attttgtaga gatattggaa agccagaagt tgaatatgat tgtgataatt tgcaacatag
      361 taagaagggg aaaactgcgc aggggttaag ccccgtggac caaaggaaat gtgaacgtct
      421 tctgctttat ctctattgcc atgaattaag tattgaattc caggagcctg ttcctgcttc
      481 gataccaaac tactataaaa ttataaagaa accaatggat ttatccaccg tgaaaaagaa
      541 gcttcagaaa aaacattccc aacactacca aatcccggat gactttgtgg ccgatgtccg
      601 tttgatcttc aagaactgtg aaaggtttaa tgaagctgat tcagaagtag ctcaggcagg
      661 gaaagcagtt gcattgtact ttgaagataa actcacagag atctactcag acaggacctt
      721 cgcacctttg ccatgacagt aaaggtggat acggatccga attcgagctc cgtcgacaag
      781 cttgcggccg cactcgagca ccaccaccac caccactgag atccggctgc taacaaagcc
      841 cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg
      901 gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc cggattggcg
      961 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     1021 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     1081 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     1141 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     1201 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     1261 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     1321 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     1381 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg
     1441 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     1501 catgaattaa ttcttagaaa aactcatcga gcatcaaatg aaactgcaat ttattcatat
     1561 caggattatc aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac
     1621 cgaggcagtt ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa
     1681 catcaataca acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac
     1741 catgagtgac gactgaatcc ggtgagaatg gcaaaagttt atgcatttct ttccagactt
     1801 gttcaacagg ccagccatta cgctcgtcat caaaatcact cgcatcaacc aaaccgttat
     1861 tcattcgtga ttgcgcctga gcgagacgaa atacgcgatc gctgttaaaa ggacaattac
     1921 aaacaggaat cgaatgcaac cggcgcagga acactgccag cgcatcaaca atattttcac
     1981 ctgaatcagg atattcttct aatacctgga atgctgtttt cccggggatc gcagtggtga
     2041 gtaaccatgc atcatcagga gtacggataa aatgcttgat ggtcggaaga ggcataaatt
     2101 ccgtcagcca gtttagtctg accatctcat ctgtaacatc attggcaacg ctacctttgc
     2161 catgtttcag aaacaactct ggcgcatcgg gcttcccata caatcgatag attgtcgcac
     2221 ctgattgccc gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg
     2281 aatttaatcg cggcctagag caagacgttt cccgttgaat atggctcata acaccccttg
     2341 tattactgtt tatgtaagca gacagtttta ttgttcatga ccaaaatccc ttaacgtgag
     2401 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     2461 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     2521 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     2581 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     2641 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     2701 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     2761 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     2821 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     2881 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     2941 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     3001 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     3061 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     3121 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     3181 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt
     3241 ctccttacgc atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct
     3301 ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc
     3361 tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     3421 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     3481 gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga
     3541 ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa
     3601 tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga
     3661 tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag
     3721 gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg
     3781 taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca
     3841 gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca
     3901 gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg
     3961 gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct
     4021 tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc
     4081 tagccgggtc ctcaacgaca ggagcacgat catgcgcacc cgtggggccg ccatgccggc
     4141 gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc
     4201 gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg
     4261 aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat
     4321 gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga
     4381 gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag
     4441 ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     4501 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca
     4561 gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc
     4621 cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt
     4681 tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta
     4741 ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg
     4801 ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca
     4861 tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa
     4921 tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac
     4981 ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc
     5041 ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga
     5101 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
     5161 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
     5221 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
     5281 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
     5341 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
     5401 tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa
     5461 cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg
     5521 cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc
     5581 gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc
     5641 tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc
     5701 accgccgccg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc cccggccacg
     5761 gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga
     5821 tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg
     5881 atgccggcca cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg aaat
//
