LOCUS       sequence-253030-        4226 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GB1 Leu5Met mutant.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4226)
  AUTHORS   Chen Q, Xu S, Lu X, Boeri MV, Pepelyayeva Y, Diaz EL, Soni SD, 
            Allaire M, Forstner MB, Bahnson BJ, Rozovsky S
  TITLE     (77)Se NMR Probes the Protein Environment of Selenomethionine.
  JOURNAL   J Phys Chem B. 2020 Jan 30;124(4):601-616. doi: 
            10.1021/acs.jpcb.9b07466. Epub 2020 Jan 7.
  PUBMED    31846581
REFERENCE   2  (bases 1 to 4226)
  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"; doi: 
            "10.1021/acs.jpcb.9b07466"; journalName: "J Phys Chem B"; date: 
            "2020-01-30- 30"; volume: "124"; issue: "4"; pages: "601-616"
COMMENT     Sequence Label: His-TEV-GB1 Leu5Met
FEATURES             Location/Qualifiers
     source          1..4226
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      86..674
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     575..594
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     828..845
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     943..960
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     944..960
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1423..1442
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1423..1441
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1442..1466
                     /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)."
     CDS             1515..1532
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1554..1574
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     terminator      1753..1800
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(2023..2042)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(2048..2067)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        2109..2200
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2201..2269,2270..3061)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2419..2438)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             3144..4226
                     /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     complement(3163..3182)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ctcatgacca aaatccctta acgtgagtta cgcgcgcgtc gttccactga gcgtcagacc
       61 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
      121 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
      181 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
      241 tgtagccgta gttagcccac cacttcaaga actctgtagc accgcctaca tacctcgctc
      301 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
      361 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
      421 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
      481 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
      541 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
      601 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
      661 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
      721 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
      781 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
      841 gcgaggaagc ggaaggcgag agtagggaac tgccaggcat caaactaagc agaaggcccc
      901 tgacggatgg cctttttgcg tttctacaaa ctctttctgt gttgtaaaac gacggccagt
      961 cttaagctcg ggccccctgg gcggttctga taacgagtaa tcgttaatcc gcaaataacg
     1021 taaaaacccg cttcggcggg tttttttatg gggggagttt agggaaagag catttgtcag
     1081 aatatttaag ggcgcctgtc actttgcttg atatatgaga attatttaac cttataaatg
     1141 agaaaaaagc aacgcacttt aaataagata cgttgctttt tcgattgatg aacacctata
     1201 attaaactat tcatctatta tttatgattt tttgtatata caatatttct agtttgttaa
     1261 agagaattaa gaaaataaat ctcgaaaata ataaagggaa aatcagtttt tgatatcaaa
     1321 attatacatg tcaacgataa tacaaaatat aatacaaact ataagatgtt atcagtattt
     1381 attatcattt agaataaatt ttgtgtcgcc cttccgcgaa attaatacga ctcactatag
     1441 gggaattgtg agcggataac aattcccctc tagaaataat tttgtttaac ttttaggagg
     1501 taaaacatat gagccatcac caccaccacc acgactacga cattccgacg acggagaact
     1561 tgtatttcca gggtcagtac aaaatgatcc tgaatggcaa gaccctgaag ggtgaaacca
     1621 ccaccgaagc ggtcgatgca gcgacggctg aaaaagtttt caagcaatat gcgaacgata
     1681 atggtgttga cggcgagtgg acttacgatg acgccacgaa aacctttacc gtgaccgagt
     1741 aactcgagcc ccctagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     1801 cccctgagac gcgtcaatcg agttcgtacc taagggcgac accccataat tagcccgggc
     1861 gaaaggccca gtctttcgac tgagcctttc gttttatttg atgcctggca gttccctact
     1921 ctcgcatggg gagtccccac actaccatcg gcgctacggc gtttcacttc tgagttcggc
     1981 atggggtcag gtgggaccac cgcgctactg ccgccaggca aacaaggggt gttatgagcc
     2041 atattcaggt ataaatgggc tcgcgataat gttcagaatt ggttaattgg ttgtaacact
     2101 gacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     2161 accctgataa atgcttcaat aatattgaaa aaggaagaat atgagtattc aacatttccg
     2221 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     2281 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     2341 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     2401 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     2461 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     2521 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     2581 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     2641 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     2701 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcga tggcaacaac
     2761 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     2821 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     2881 gtttattgct gataaatccg gagccggtga gcgtggttct cgcggtatca tcgcagcgct
     2941 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     3001 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     3061 agcggcgcgc catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag
     3121 agagtcaatt cagggtggtg aatatgaaac cagtaacgtt atacgatgtc gcagagtatg
     3181 ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga
     3241 aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg
     3301 cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc
     3361 tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca
     3421 gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca
     3481 atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg
     3541 ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc
     3601 agacacccat caacagtatt attttctccc atgaggacgg tacgcgactg ggcgtggagc
     3661 atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct
     3721 cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa attcagccga
     3781 tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc
     3841 tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg gcgctgggcg
     3901 caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatctcg gtagtgggat
     3961 acgacgatac cgaagatagc tcatgttata tcccgccgtt aaccaccatc aaacaggatt
     4021 ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag ggccaggcgg
     4081 tgaagggcaa tcagctgttg ccagtctcac tggtgaaaag aaaaaccacc ctggcgccca
     4141 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
     4201 tttcccgact ggaaagcggg cagtga
//
