LOCUS       sequence-217475-        7019 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For protein expression and purification of Drosophila CvcE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7019)
  TITLE     Orr-Weaver plasmids
REFERENCE   2  (bases 1 to 7019)
  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"
COMMENT     Sequence Label: CvcE in pGEX4t
FEATURES             Location/Qualifiers
     source          1..7019
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     154..171
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        244..321
                     /gene="lacI"
                     /label=lacI promoter
     CDS             322..1404
                     /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(341..360)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1453..1458,1459..1476,1477..1483)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1491..1507
                     /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     1515..1531
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1515..1531
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1527..2054
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(1547..1564)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1547..1563)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1556..1578)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(2156..2161,2162..2177,2178..2184)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2192..2208
                     /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             2231..2884
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2842..2864
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2891..2908
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     protein_bind    3368..3379
                     /label=ZF binding site
                     /bound_moiety="CCR5TC zinc-finger domain"
                     /note="target sequence for the CCR5TC zinc-finger domain 
                     from the CCR5-224 (+) zinc-finger nuclease (Perez et al., 
                     2008; Gross et al., 2013)"
     primer_bind     complement(5042..5064)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5209..5227)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5295..5399
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5400..5468,5469..6260)
                     /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(5618..5637)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6431..7019
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6920..6939
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
       61 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
      121 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      181 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcataaat
      241 tccgacacca tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag
      301 agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc
      361 ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa
      421 acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca
      481 caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg
      541 cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc
      601 gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat
      661 cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc
      721 attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag
      781 acacccatca acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat
      841 ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg
      901 gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata
      961 gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg
     1021 aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca
     1081 atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac
     1141 gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt
     1201 cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg
     1261 aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat
     1321 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     1381 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta
     1441 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     1501 ataacaattt cacacaggaa acagctatga ccatgattac ggattcactg gccgtcgttt
     1561 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
     1621 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
     1681 tgcgcagcct gaatggcgaa tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg
     1741 aaagctggct ggagtgcgat cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc
     1801 agatgcacgg ttacgatgcg cccatctaca ccaacgtgac ctatcccatt acggtcaatc
     1861 cgccgtttgt tcccacggag aatccgacgg gttgttactc gctcacattt aatgttgatg
     1921 aaagctggct acaggaaggc cagacgcgaa ttatttttga tggcgttgga attagcttat
     1981 cgactgcacg gtgcaccaat gcttctggcg tcaggcagcc atcggaagct gtggtatggc
     2041 tgtgcaggtc gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc cgttctggat
     2101 aatgtttttt gcgccgacat cataacggtt ctggcaaata ttctgaaatg agctgttgac
     2161 aattaatcat cggctcgtat aatgtgtgga attgtgagcg gataacaatt tcacacagga
     2221 aacagtattc atgtccccta tactaggtta ttggaaaatt aagggccttg tgcaacccac
     2281 tcgacttctt ttggaatatc ttgaagaaaa atatgaagag catttgtatg agcgcgatga
     2341 aggtgataaa tggcgaaaca aaaagtttga attgggtttg gagtttccca atcttcctta
     2401 ttatattgat ggtgatgtta aattaacaca gtctatggcc atcatacgtt atatagctga
     2461 caagcacaac atgttgggtg gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg
     2521 agcggttttg gatattagat acggtgtttc gagaattgca tatagtaaag actttgaaac
     2581 tctcaaagtt gattttctta gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt
     2641 atgtcataaa acatatttaa atggtgatca tgtaacccat cctgacttca tgttgtatga
     2701 cgctcttgat gttgttttat acatggaccc aatgtgcctg gatgcgttcc caaaattagt
     2761 ttgttttaaa aaacgtattg aagctatccc acaaattgat aagtacttga aatccagcaa
     2821 gtatatagca tggcctttgc agggctggca agccacgttt ggtggtggcg accatcctcc
     2881 aaaatcggat ctggttccgc gtggatcctg ttattgtctt gagtatcctc tccactcatt
     2941 tgttcatgtt attatgaagt tggaacagaa gcgcaaattt attgaaatgg accctgaact
     3001 cggttttgag cctccatcag ctaagcgcca gcagcgactg cccgctttgt acggcagcga
     3061 gcagggcaat ctgtcatcgg ttgcctcctc ggtctacacc tcgcccgttg tctcggttga
     3121 tggccagagc acccaggagc tgctcagcat acgtagctca ccggccgagg atctgtcgga
     3181 ggcgccacac agtccgctgc cggacagccc ggacagtccg ccgagtccgg atcgtggaag
     3241 caagcagacg ccagtggtgg tgcgctacgc tgcggagcag gtggtgacca gtacggtggt
     3301 cacgcagaag acggaggatg atgacctttt ggacgacagc tgcgaagact actcgtacga
     3361 cgaggatgat gaggatgacg ttgaggagga ggacgacgac gtggagatct actcctccac
     3421 aatttctcca gcctcctccg gctgcagcca gcagcaggcg gtgaatggag agcgtactcc
     3481 cggcctgcca aaacatcagg agcagatcca tcatccggtc agtgacctga tgattaacat
     3541 gcggacgccc atgtcgccgg ctgtggagaa cggtctgcgc cagtgcccac ttcctgcgct
     3601 cgcctgggcc aacgccgccg atgtttggcg cctgatgtgt caccgggatg agcaggactc
     3661 gcgtctgcgc agcatttcga tgctggaaca gcatcccgga ctgcaaccac gtatgcgtgc
     3721 cattctcttg gactggttga tcgaggtctg tgaagtttac aagctgcatc gggagacctt
     3781 ctacttggcc gtcgactacc tggatcgcta tttgcatgtg gcgcacaagg tgcagaagac
     3841 gcacttgcag ttgatcggca tcacctgcct gtttgtggcc gccaaggtag aggagattta
     3901 tccgccaaag atcggagagt ttgcttatgt gacggatggt gcctgcacag agcgcgacat
     3961 ccttaaccac gagaagattc tgctgcaggc gctcgactgg gacatcagcc ccatcaccat
     4021 taccggctgg ctgggcgtct atatgcaact gaatgtgaac aaccgcactc cggcctcgtt
     4081 ctcccagatt ggcaggcaaa agagcgcgga ggcggacgat gcctttatct acccgcaatt
     4141 ctctggcttt gagtttgtgc aaacctcaca gctgctagat ctgtgcaccc tggacgtggg
     4201 catggccaat tactcctact cggtgctggc agcggccgcc atcagtcata catttagtcg
     4261 ggagatggcc ttacgctgtt ctggtctgga ttggcaggtg attcagcctt gtgctcgctg
     4321 gatggagcca ttcttccgag tgatctccca gaaggccccc tacctgcagc tgaacgagca
     4381 gaatgagcaa gtcagcaaca aatttggcct gggactaatc tgccccaata ttgtcaccga
     4441 cgactcgcac attatccaaa cgcacacaac taccatggat atgtatgacg aagtgcttat
     4501 ggctcaggat gcagcgcatg ccatgcgctc taggattcaa gcttccccgg ccaccgcgct
     4561 gcgcgctccc gaaagtcttc tgacgcctcc cgcctctagt cacaagccgg acgagtacct
     4621 gggcgatgag ggcgatgaga cgggggccag gagcggcatc tcctccacaa ccacatgctg
     4681 caacactgcc gccagtaaca aggtgggcaa gagcagcagc aacaattcgg tgaccagctg
     4741 cagcagtaga agcaatccct gagagattcg cctcactgct cttggccatc atcctctcac
     4801 actcaccaca cccgcccaaa cagtctacgt taagtgtttc ttatatttaa attatttttg
     4861 ttaagtgaaa agttattagt tgcctaattc ctatgctaca aacatattta atttgaaatt
     4921 atctacgcgc tcttgattct caactcgcga tataatcatt ggaattcccg ggtcgactcg
     4981 agcggccgca tcgtgactga ctgacgatct gcctcgcgcg tttcggtgat gacggtgaaa
     5041 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     5101 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga
     5161 cccagtcacg tagcgatagc ggagtgtata attcttgaag acgaaagggc ctcgtgatac
     5221 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     5281 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     5341 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     5401 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5461 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     5521 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     5581 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     5641 gtgttgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     5701 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     5761 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     5821 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     5881 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     5941 ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     6001 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     6061 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     6121 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     6181 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     6241 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     6301 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     6361 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     6421 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     6481 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     6541 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     6601 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     6661 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     6721 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     6781 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     6841 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     6901 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     6961 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaa
//
