LOCUS       sequence-305914-        6707 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To express LanCL2 in bacterial cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6707)
  AUTHORS   Lai KY, Galan SRG, Zeng Y, Zhou TH, He C, Raj R, Riedl J, Liu S, 
            Chooi KP, Garg N, Zeng M, Jones LH, Hutchings GJ, Mohammed S, Nair 
            SK, Chen J, Davis BG, van der Donk WA
  TITLE     LanCLs add glutathione to dehydroamino acids generated at 
            phosphorylated sites in the proteome.
  JOURNAL   Cell. 2021 Apr 27. pii: S0092-8674(21)00436-0. doi: 
            10.1016/j.cell.2021.04.001.
  PUBMED    33932340
REFERENCE   2  (bases 1 to 6707)
  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"; journalName: "Cell. 2021 
            Apr 27. pii: S0092-8674(21)00436-0. doi: 
            10.1016/j.cell.2021.04.001."
COMMENT     Sequence Label: pETDuet1-WT-LanCL2-C-Ter-His
FEATURES             Location/Qualifiers
     source          1..6707
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     90..109
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        197..215
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    216..240
                     /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             255..277
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1634..1651
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        1717..1735
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1736..1760
                     /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             1869..1913
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1951..1969)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1965..2012
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2049..2504
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2136..2155)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2346..2367
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(2619..3410,3411..3479))
                     /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     3242..3261
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3480..3572)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3653..4241
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4142..4161
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4395..4412
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4427..4566
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4492..4511)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4652..4674
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4668..4859)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5431..6513)
                     /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     6475..6494
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6514..6591)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
       61 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
      121 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat cgagatcgat
      181 ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga taacaattcc
      241 cctctagaaa taattttgtt taactttaag aaggagatat accatgggcg agaccatgtc
      301 aaagaggctg aagctccacc tgggagggga ggcagaaatg gaggaacggg cgttcgtcaa
      361 ccccttcccg gactacgagg ccgccgccgg ggcgctgctc gcctccggag cggccgaaga
      421 gacaggctgt gttcgtcccc cggccacccc ggatgagccc ggcctccctt ttcatcagga
      481 cgggaagatc attcataatt tcataagacg gatccagacc aaaattaaag atcttctgca
      541 gcaaatggaa gaagggctga agacagctga tccccatgac tgctctgctt atactggctg
      601 gacaggcata gcccttttgt acctgcagtt gtaccgggtc acatgtgacc aaacctacct
      661 gctccgatcc ctggattacg taaaaagaac acttcggaat ctgaatggcc gcagggtcac
      721 cttcctctgt ggggatgctg gccccctggc tgttggagct gtgatttatc acaaactcag
      781 aagtgactgt gagtcccagg aatgtgtcac aaaacttttg cagctccaga gatcggttgt
      841 ctgccaagaa tcagaccttc ctgatgagct gctttatgga cgggcaggtt atctgtatgc
      901 cttactgtac ctgaacacag agataggtcc aggcaccgtg tgtgagtcag ctattaaaga
      961 ggtagtcaat gctattattg aatcgggtaa gactttgtca agggaagaaa gaaaaacgga
     1021 gcgctgcccg ctgttgtacc agtggcaccg gaagcagtac gttggagcag cccatggcat
     1081 ggctggaatt tactatatgt taatgcagcc ggcagcaaaa gtggaccaag aaaccttgac
     1141 agaaatggtg aaacccagta ttgattatgt gcgccacaaa aaattccgat ctgggaatta
     1201 cccatcatca ttaagcaatg aaacagaccg gctggtgcac tggtgccacg gcgccccggg
     1261 ggtcatccac atgctcatgc aggcgtacaa ggtctttaag gaggagaagt acttgaaaga
     1321 ggccatggag tgtagcgatg tgatttggca gcgaggtttg ctgcggaagg gctacgggat
     1381 atgccatggg actgctggca acggctattc cttcctgtcc ctttaccgtc tcacgcagga
     1441 taagaagtac ctctaccgag cttgcaagtt tgcagagtgg tgtctagatt acggagcaca
     1501 cgggtgccgc attcctgaca gaccctattc gctctttgaa ggcatggctg gcgctattca
     1561 ctttctctct gatgtcctgg gaccagagac atcacggttt ccagcatttg aacttgactc
     1621 ttcgaagagg gatcatcacc atcatcacca cgcggccgca taatgcttaa gtcgaacaga
     1681 aagtaatcgt attgtacacg gccgcataat cgaaattaat acgactcact ataggggaat
     1741 tgtgagcgga taacaattcc ccatcttagt atattagtta agtataagaa ggagatatac
     1801 atatggcaga tctcaattgg atatcggccg gccacgcgat cgctgacgtc ggtaccctcg
     1861 agtctggtaa agaaaccgct gctgcgaaat ttgaacgcca gcacatggac tcgtctacta
     1921 gcgcagctta attaacctag gctgctgcca ccgctgagca ataactagca taaccccttg
     1981 gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata tccggattgg
     2041 cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     2101 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     2161 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     2221 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     2281 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     2341 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     2401 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     2461 aaaatttaac gcgaatttta acaaaatatt aacgtttaca atttctggcg gcacgatggc
     2521 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     2581 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     2641 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     2701 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     2761 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     2821 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     2881 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     2941 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     3001 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     3061 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     3121 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     3181 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     3241 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     3301 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     3361 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     3421 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     3481 ctcttccttt ttcaatcatg attgaagcat ttatcagggt tattgtctca tgagcggata
     3541 catatttgaa tgtatttaga aaaataaaca aataggtcat gaccaaaatc ccttaacgtg
     3601 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     3661 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     3721 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     3781 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     3841 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     3901 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     3961 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     4021 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     4081 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     4141 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     4201 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     4261 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     4321 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     4381 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt
     4441 ttctccttac gcatctgtgc ggtatttcac accgcaatgg tgcactctca gtacaatctg
     4501 ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg
     4561 gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     4621 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     4681 ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc
     4741 gattcacaga tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt
     4801 aatgtctggc ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact
     4861 gatgcctccg tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag
     4921 aggatgctca cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag
     4981 ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc
     5041 cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg
     5101 cagatccgga acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca
     5161 cggaaaccga agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg
     5221 cttcacgttc gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag
     5281 cctagccggg tcctcaacga caggagcacg atcatgctag tcatgccccg cgcccaccgg
     5341 aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg tgcctaatga
     5401 gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     5461 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     5521 cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc ccttcaccgc
     5581 ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc
     5641 ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat cgtcgtatcc
     5701 cactaccgag atgtccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc
     5761 cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat
     5821 ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg
     5881 ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac
     5941 agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc
     6001 cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc
     6061 agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc
     6121 ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg
     6181 caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc
     6241 acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc
     6301 cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac
     6361 gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc
     6421 agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata
     6481 ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc
     6541 cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc
     6601 gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt
     6661 tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgccc
//
