LOCUS       sequence-462744-        6919 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial expression of His-tagged GNL2 (1-645aa). To be used as a 
            probe..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6919)
  TITLE     Iuchi Lab GNL2 Plasmids 2025
REFERENCE   2  (bases 1 to 6919)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pGNL2-#21
FEATURES             Location/Qualifiers
     source          1..6919
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             921..932
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             2883..2900
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2969..2991)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3136..3154)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3222..3326
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3327..3395,3396..4187)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3545..3564)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4358..4946
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4847..4866
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5100..5117
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        5190..5267
                     /gene="lacI"
                     /label=lacI promoter
     CDS             5268..6350
                     /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(5287..5306)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(6399..6404,6405..6422,6423..6429)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6437..6453
                     /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)."
     primer_bind     6461..6477
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6461..6477
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(6473..6919,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6493..6510)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6493..6509)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6502..6524)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg atcgaaggtc gtgggatccc cgaattcatg gtgaagccca
      961 agtacaaagg acggagcacc atcaacccgt ccaaggccag cacaaaccca gatcgagtgc
     1021 agggagcagg aggccaaaac atgagggacc gggccaccat ccggcgcctg aatatgtata
     1081 ggcaaaagga gcgcaggaac agtcgtggta aaataattaa acccctgcaa tatcaatcaa
     1141 cggtggcttc tggcacagtg gcaagagtag agccaaatat taaatggttt ggaaacacac
     1201 gtgtgattaa gcagtcatca ttacaaaaat ttcaagagga aatggataca gttatgaagg
     1261 atccatacaa agttgtcatg aagcaaagca agttaccaat gtctcttctc catgatcgaa
     1321 tccggcctca taacttgaag gtgcacattc ttgatactga aagttttgaa actacatttg
     1381 gccctaagtc acagaggaaa cgaccaaact tatttgcaag tgatatgcag tctcttatcg
     1441 aaaatgctga aatgtccact gagagctatg accagggcaa ggatcgtgat ttggtaactg
     1501 aagacactgg tgtgagaaat gaagctcaag aagagatcta taaaaaggga cagtccaaaa
     1561 gaatatgggg tgagctctac aaggtgatag attcatcaga tgttgtagtt caagttcttg
     1621 atgctagaga tccaatgggt actcgttccc ctcacattga aacttacctg aagaaggaaa
     1681 aaccttggaa acacctcatt tttgtactta acaaatgtga ccttgttcca acctgggcaa
     1741 caaaacggtg ggttgctgtc ctctcccagg attatccaac acttgctttc catgcaagcc
     1801 ttactaaccc gtttggcaag ggagcattca ttcagcttct gcggcagttt ggaaagttgc
     1861 acactgacaa gaaacagatc agtgttgggt tcattggcta tccaaatgtt ggcaagagct
     1921 ctgtgataaa tacattgcgt tctaagaaag tttgcaacgt ggctcccatt gcaggtgaaa
     1981 caaaggtctg gcagtatatt actttgatgc gtcggatatt cctgattgac tgtccaggtg
     2041 tggtttaccc ctctgaggac tccgagacag acattgtgct aaaaggagtt gttcaagtag
     2101 aaaaaattaa gagtcctgaa gaccacattg gtgctgtact tgaacgagca aagccagaat
     2161 atatcagcaa aacatacaag attgattctt gggagaatgc tgaggacttt cttgagaagc
     2221 tcgctttccg gactgggaag ttactaaagg gtggagagcc cgacttgcag actgtgggta
     2281 agatggtcct caatgactgg cagaggggcc ggattccttt ctttgtcaag ccacccaatg
     2341 cagagccact tgtggccccc cagcttctac cctcctcatc tttggaagtt gtcccagaag
     2401 cagcccagaa caatccaggg gaggaagtca cagaaactgc aggtgaaggg tcagaatcca
     2461 tcattaagga agaaacagaa gagaacagtc actgtgatgc taacacagag atgcagcaga
     2521 ttctcacacg agttcggcag aactttggta aaatcaacgt ggtgcctcag ttttctgggg
     2581 atgacctggt tcctgtggag gtgtcagatc ttgaggaaga gcttgagagc ttttctgatg
     2641 aagaggagga ggaacaggag caacaaagag atgatgcgga agagtcttcc tcggagcctg
     2701 aggaggaaaa tgtgggaaac gacaccaaag ccgttattaa agcactggat gagaagattg
     2761 ccaaatatca gaagtttcta gacaaagcca aagccaaaaa gttttcagca gtcagaatat
     2821 ccaagggact gagtgaaaag atatttgcaa aacctgaaga acaaagaaaa acactggaag
     2881 aacaccacca ccaccaccac tgactcgagc ggccgcatcg tgactgactg acgatctgcc
     2941 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
     3001 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
     3061 ttggcgggtg tcggggcgca gccatgaccc agtcacgtag cgatagcgga gtgtataatt
     3121 cttgaagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     3181 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     3241 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     3301 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     3361 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     3421 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     3481 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     3541 ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc
     3601 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     3661 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     3721 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     3781 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     3841 caaacgacga gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat
     3901 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     3961 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     4021 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     4081 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     4141 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     4201 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     4261 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     4321 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     4381 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     4441 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     4501 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     4561 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     4621 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     4681 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4741 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4801 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4861 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     4921 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     4981 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     5041 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     5101 gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc
     5161 tgtgcggtat ttcacaccgc ataaattccg acaccatcga atggcgcaaa acctttcgcg
     5221 gtatggcatg atagcgcccg gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa
     5281 cgttatacga tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga
     5341 accaggccag ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc
     5401 tgaattacat tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg
     5461 gcgttgccac ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat
     5521 ctcgcgccga tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg
     5581 aagcctgtaa agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta
     5641 actatccgct ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg
     5701 cgttatttct tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgaag
     5761 acggtacgcg actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt
     5821 tagcgggccc attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc
     5881 tcactcgcaa tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg
     5941 gttttcaaca aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg
     6001 ccaacgatca gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg
     6061 gtgcggatat ctcggtagtg ggatacgacg ataccgaaga cagctcatgt tatatcccgc
     6121 cgttaaccac catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc
     6181 tgcaactctc tcagggccag gcggtgaagg gcaatcagct gttgcccgtc tcactggtga
     6241 aaagaaaaac caccctggcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     6301 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
     6361 attaatgtaa gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct
     6421 cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat
     6481 gattacggat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     6541 ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc
     6601 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc gctttgcctg
     6661 gtttccggca ccagaagcgg tgccggaaag ctggctggag tgcgatcttc ctgaggccga
     6721 tactgtcgtc gtcccctcaa actggcagat gcacggttac gatgcgccca tctacaccaa
     6781 cgtgacctat cccattacgg tcaatccgcc gtttgttccc acggagaatc cgacgggttg
     6841 ttactcgctc acatttaatg ttgatgaaag ctggctacag gaaggccaga cgcgaattat
     6901 ttttgatggc gttggaatt
//
