LOCUS       sequence-444363-        6513 bp    DNA     circular SYN 15-APR-2025
DEFINITION  IPTG-inducible expression of C. elegans protein ADH-5 with a His 
            tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6513)
  AUTHORS   Katherine Walstrom Lab
  TITLE     Walstrom Lab Plasmids 2024
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6513)
  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"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: ADH5_KG
FEATURES             Location/Qualifiers
     source          1..6513
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      28..616
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     517..536
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     770..787
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    802..941
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(867..886)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1027..1049
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1043..1234)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2043..3125)
                     /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     3087..3106
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3126..3203)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3409..3428
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3512..3531
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3512..3530
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3531..3555
                     /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             3571..3591
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4756..4773
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4826..4844)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4840..4887
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4924..5379
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5011..5030)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5221..5242
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5405..5509
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5510..5578,5579..6370)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5728..5747)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
       61 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
      121 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
      181 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
      241 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
      301 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
      361 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
      421 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
      481 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
      541 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
      601 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
      661 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
      721 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
      781 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
      841 ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
      901 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
      961 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     1021 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     1081 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
     1141 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
     1201 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
     1261 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
     1321 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
     1381 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
     1441 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
     1501 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
     1561 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
     1621 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
     1681 gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga
     1741 aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata
     1801 ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga
     1861 cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg
     1921 cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca
     1981 agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc
     2041 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     2101 aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt
     2161 gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg
     2221 tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata
     2281 taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc
     2341 agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc
     2401 atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg
     2461 gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta
     2521 tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg
     2581 atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg
     2641 gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca
     2701 ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc
     2761 agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg
     2821 cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc
     2881 gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc
     2941 aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc
     3001 atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg
     3061 cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt
     3121 ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag
     3181 gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat
     3241 taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc
     3301 atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc
     3361 gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc
     3421 gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc
     3481 gtagaggatc gagatctcga tcccgcgaaa ttaatacgac tcactatagg ggaattgtga
     3541 gcggataaca attcccctct agtaataatt ttgtttaact ttaagaagga ggtctagatg
     3601 agctccaccg ctggacaggt tatcaactgc aaggcagccg tcgcctggag cgcaaaagct
     3661 ccactctcga ttgagacgat ccaagtggct ccaccgaaag cccacgaagt tcgagtgaaa
     3721 atcctctaca ccgcagtctg ccacacggac gcctacaccc tcgacggaca cgatccggaa
     3781 ggtctcttcc ctgtcgtctt gggacatgaa gggtctggaa ttgtcgagag cgtcggagaa
     3841 ggagtcacag gatttgctcc aggagatcac gtcgttccgc tctacgttcc tcagtgcaaa
     3901 gaatgtgaat attgcaagaa tccaaagacg aatctgtgtc aaaagatcag aatctcgcag
     3961 ggaaatggat ttatgcccga tggttcatcg agattcacat gcaacggaaa gcagttgttc
     4021 cattttatgg gatgctccac gttcagtgaa tacactgttg tggccgacat ctcactgtgc
     4081 aaagtgaatc cagaagctcc gctcgagaag gtttcgcttt tgggatgtgg aatttcaact
     4141 ggatacggag ccgtgctgaa cacgtgcaaa gtggaagaag ggagtactgt agccgtatgg
     4201 ggactcggtg cagtcggatt ggccgttatc atgggcgcaa aggctgccgg agccaagaaa
     4261 atcgtcggaa tcgacctgat cgagtcgaaa ttcgagtcgg ccaaattctt cggagccacc
     4321 gaatgcatca atccgaagag tgttgagctt cctgagggga agagcttcca agcatggctc
     4381 gtggagcaat ttgacggagg atttgactac acttttgagt gtattggaaa tgtgcacaca
     4441 atgcgccaag ccctcgaagc cgcccacaaa ggatggggag tctcgtgtat catcggagtc
     4501 gccggagccg gccaagagat cgccacccgt ccattccaac ttgtcaccgg gcgcacgtgg
     4561 aagggaaccg cattcggcgg atggaaatcc gtggaaagtg tgccgagact cgtcgacgac
     4621 tacatgaaca agaagctgct catcgatgaa ttcattactc atcgctggaa cattgacgat
     4681 atcaatacgg cgttcgatgt tcttcacaag ggagagagtc tgcggtcggt gctggcgttc
     4741 gagaaaatac tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga
     4801 aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc
     4861 tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg attggcgaat
     4921 gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     4981 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     5041 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     5101 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     5161 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     5221 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     5281 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     5341 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa
     5401 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     5461 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     5521 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     5581 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     5641 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     5701 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     5761 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     5821 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     5881 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     5941 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     6001 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat
     6061 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     6121 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     6181 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     6241 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     6301 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     6361 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     6421 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     6481 ttaacgtgag ttttcgttcc actgagcgtc aga
//
