LOCUS       sequence-288828-        6245 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of AfPiwi.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6245)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 6245)
  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: pGEX-CS-AfPiwi_G
FEATURES             Location/Qualifiers
     source          1..6245
                     /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)."
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     primer_bind     complement(2295..2317)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2462..2480)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2548..2652
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2653..2721,2722..3513)
                     /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(2871..2890)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3684..4272
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4173..4192
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4426..4443
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4516..4593
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             4594..5676
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4613..4632)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    5689..5710
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5725..5730,5731..5748,5749..5755)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5763..5779
                     /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     5787..5803
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5787..5803
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5799..6245,1..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(5819..5836)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5819..5835)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5828..5850)
                     /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 atctcgagat ctttatttta gggcgccatg gcaatgatgg aatataaaat agttgaaaat
      961 ggtttgacat accgtatagg gaatggcgca tctgtaccaa tatcaaacac tggtgagttg
     1021 attaaaggat taaggaatta tgggccttat gaagtcccct ccttaaagta taatcaaatt
     1081 gcattaatcc ataacaatca gttttcaagt ttaataaacc aattgaaatc acaaataagt
     1141 tctaaaattg atgaggtgtg gcatattcat aatataaaca tttctgagtt tatctacgat
     1201 tccccacact ttgactctat caaatctcaa gtagataatg ctatagatac gggtgtggac
     1261 ggaattatgc tcgttcttcc aggatataat accccattat attataagct taaatcgtat
     1321 ttaataaata gtatcccatc acaatttatg agatatgata tattatctaa cagaaatttg
     1381 acattctatg ttgacaatct gttggtacaa tttgtttcaa aacttggtgg aaaaccttgg
     1441 attttaaatg tagatcctga aaaaggttca gacattataa taggaactgg cgcaacacgt
     1501 attgataatg taaatctatt ttgttttgct atggtcttca agaaagatgg gacaatgctg
     1561 tggaatgaaa tttcaccaat agttacgagt agtgaatatc taacatattt aaaaagtaca
     1621 attaagaaag tggtatatgg tttcaaaaaa agtaatccgg attgggatgt cgaaaagtta
     1681 actttacatg taagtggaaa aaggcccaaa atgaaagatg gagagacaaa gattttaaaa
     1741 gaaacggttg aagagttaaa aaaacaagaa atggtatcaa gagacgtaaa atatgctata
     1801 cttcgcttaa atgaaactca tccattttgg gtaatgggag accctaacaa tagatttcat
     1861 ccgtatgaag ggacaaaagt aaaattaagt tctaaaagat acttattaac acttttacag
     1921 ccttatctta aacgaaatgg attagaaatg gtaaccccta taaaaccatt aagtgttgag
     1981 atcgttagcg ataattggac ttctgaagaa tactaccaca atgttcatga gattttggat
     2041 gaaatatatt atctgtctaa aatgaattgg agaggattta gaagtagaaa tctacccgtt
     2101 acggttaatt atccaaaatt ggtcgcagga atcatagcaa atgtaaatcg ttatggagga
     2161 tatccaataa acccagaagg gaatcgctca ttacaaacga atccgtggtt cctatgaggt
     2221 acccggggat ccccgggaat tcatcgtgac tgactgacga tctgcctcgc gcgtttcggt
     2281 gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa
     2341 gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg
     2401 ggcgcagcca tgacccagtc acgtagcgat agcggagtgt ataattcttg aagacgaaag
     2461 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     2521 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     2581 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     2641 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     2701 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     2761 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     2821 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     2881 gcggtattat cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     2941 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     3001 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     3061 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     3121 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     3181 gacaccacga tgcctgcagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     3241 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     3301 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     3361 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     3421 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     3481 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     3541 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     3601 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     3661 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     3721 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     3781 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     3841 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     3901 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     3961 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     4021 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     4081 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     4141 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     4201 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     4261 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     4321 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     4381 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     4441 gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     4501 caccgcataa attccgacac catcgaatgg tgcaaaacct ttcgcggtat ggcatgatag
     4561 cgcccggaag agagtcaatt cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc
     4621 gcagagtatg ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac
     4681 gtttctgcga aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc
     4741 aaccgcgtgg cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc
     4801 agtctggccc tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa
     4861 ctgggtgcca gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg
     4921 gcggtgcaca atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat
     4981 gaccaggatg ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat
     5041 gtctctgacc agacacccat caacagtatt attttctccc atgaagacgg tacgcgactg
     5101 ggcgtggagc atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta
     5161 agttctgtct cggcgcgtct gcgtctggct ggctggcata aatatctcac tcgcaatcaa
     5221 attcagccga tagcggaacg ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc
     5281 atgcaaatgc tgaatgaggg catcgttccc actgcgatgc tggttgccaa cgatcagatg
     5341 gcgctgggcg caatgcgcgc cattaccgag tccgggctgc gcgttggtgc ggatatctcg
     5401 gtagtgggat acgacgatac cgaagacagc tcatgttata tcccgccgtc aaccaccatc
     5461 aaacaggatt ttcgcctgct ggggcaaacc agcgtggacc gcttgctgca actctctcag
     5521 ggccaggcgg tgaagggcaa tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc
     5581 ctggcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
     5641 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
     5701 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
     5761 aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt acggattcac
     5821 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     5881 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     5941 cttcccaaca gttgcgcagc ctgaatggcg aatggcgctt tgcctggttt ccggcaccag
     6001 aagcggtgcc ggaaagctgg ctggagtgcg atcttcctga ggccgatact gtcgtcgtcc
     6061 cctcaaactg gcagatgcac ggttacgatg cgcccatcta caccaacgta acctatccca
     6121 ttacggtcaa tccgccgttt gttcccacgg agaatccgac gggttgttac tcgctcacat
     6181 ttaatgttga tgaaagctgg ctacaggaag gccagacgcg aattattttt gatggcgttg
     6241 gaatt
//
