LOCUS       sequence-288831-        7458 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial Expression of SpAgo1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7458)
  TITLE     Izaurralde lab plasmids
REFERENCE   2  (bases 1 to 7458)
  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-SpAgo1_G
FEATURES             Location/Qualifiers
     source          1..7458
                     /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(3508..3530)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3675..3693)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3761..3865
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3866..3934,3935..4726)
                     /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(4084..4103)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4897..5485
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5386..5405
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5639..5656
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        5729..5806
                     /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             5807..6889
                     /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(5826..5845)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    6902..6923
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6938..6943,6944..6961,6962..6968)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6976..6992
                     /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     7000..7016
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7000..7016
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(7012..7458,1..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(7032..7049)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7032..7048)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7041..7063)
                     /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 atctcgagaa tctttatttt agggcgccat ggcaatgtcg tataaaccaa gctcagaaat
      961 agctttacgt cccggttatg gagggttggg caagcagatt actttgaaag ctaacttttt
     1021 tcaaatcatt tctttaccta acgagactat caaccagtat catgtcatag taggtgatgg
     1081 ttcacgtgtc ccacgaaagc aatcccagtt gatttggaat tctaaagaag ttaaacagta
     1141 ttttggatct tcctggatga attctgttta tgatgggcga agcatgtgtt ggtctaaagg
     1201 tgatattgct gatggcacca tcaaggtaaa cataggcagt gagtcgcacc ctcgagaaat
     1261 tgaattctct attcaaaaat catctaaaat taatttgcat actcttagcc agtttgtaaa
     1321 ctcaaagtat tcttcggatc cgcaagttct aagcagtata atgtttctgg atttgctgtt
     1381 aaagaagaaa ccatctgaga ctttgtttgg cttcatgcat tcttttttta caggtgaaaa
     1441 cggagtttca cttggaggtg gagtagaggc ttggaaaggg ttttaccagt ctatacgacc
     1501 gaatcagggt ttcatgtctg ttaatgtaga tatatcttcg agtgcatttt ggcgtaatga
     1561 ttctttactt cagatcctta tggaatacac agattgctcg aacgttagag acttaacacg
     1621 ttttgaccta aaacgtcttt cccgcaagtt tcggtttcta aaggtgacct gtcagcatcg
     1681 caataatgtc ggtactgacc tggcgaatag agtttattcc atcgagggat tttcctcaaa
     1741 aagtgccagt gatagctttt ttgttcgccg tcttaatggt gaagagcaaa aaatctctgt
     1801 agctgaatac tttcttgaaa atcacaatgt tcgtttgcaa tatcctaacc taccttgcat
     1861 attagtaaag aacggggcga tgcttcctat tgaattctgt tttgttgtaa aaggtcagag
     1921 atatactgca aaactgaata gtgatcaaac tgccaacatg atccgttttg cggtccaacg
     1981 acctttcgaa cgtgtccagc aaatcgatga ttttgtccat cagatggatt gggatacaga
     2041 tccctaccta acccaatatg gtatgaaaat tcagaaaaag atgctcgaag ttccagctcg
     2101 tgttttagaa actccttcta ttcgttacgg aggagattgc attgagcgtc ccgttagtgg
     2161 acgttggaat ttacgtggta aacggttcct tgatccgccg agggctccta ttagatcttg
     2221 ggctgtaatg tgttttactt caacgcgtag actccctatg aggggtattg aaaatttcct
     2281 gcaaacatat gtacaaacac tcacttcgtt gggaattaat tttgttatga aaaaacctcc
     2341 agttttgtat gcagatatta gagggagtgt tgaagaacta tgtataacgt tatataaaaa
     2401 agctgaacag gttggtaatg ctcctcctga ctatctcttt tttattttgg ataaaaattc
     2461 gcctgaacca tatggttcca ttaaacgtgt ttgcaatact atgttagggg ttccttctca
     2521 atgtgctatt agcaaacata ttttacaaag caagccccaa tactgtgcaa acttaggcat
     2581 gaagattaat gtcaaggttg gaggaattaa ttgctcgttg atacccaaat ccaacccttt
     2641 gggaaatgtt cctactctta ttcttggtgg agatgtttat caccctgggg ttggtgcaac
     2701 aggtgtttcg attgcttcaa ttgtggcatc ggtagaccta aatggctgca agtatacggc
     2761 tgtttcacgt tcccaacctc gtcatcaaga agtgattgaa ggaatgaagg atatcgtggt
     2821 atatctattg caagggttta gggcaatgac taaacaacaa ccacaacgta ttatctattt
     2881 ccgtgacggt acctcggaag gacaattttt atcagtaata aacgatgagc tttctcagat
     2941 taaagaagcg tgtcatagtc tatcaccaaa atacaatcca aagattctag tatgcacaac
     3001 gcagaagaga catcacgctc gtttcttcat taagaacaaa tccgacggag atagaaatgg
     3061 aaatcccctc cctggtacaa ttattgaaaa gcatgttact cacccctatc agtacgattt
     3121 ttaccttata tcccacccat ctctgcaagg cgtttcggtc ccagttcact acacggtttt
     3181 gcacgacgaa attcagatgc caccggacca attccaaaca ctgtgttata atctttgtta
     3241 tgtttatgca agagcaacaa gtgctgtttc cttggttcca cctgtttatt atgcgcattt
     3301 ggttagtaac ttagcccgtt atcaagacgt gacagccgat gatacttttg ttgagacatc
     3361 agaagcatca atggatcaag aagtaaagcc gcttttagct ctatcaagta aattgaaaac
     3421 aaagatgtgg tatatgtgag gatccccggg aattcatcgt gactgactga cgatctgcct
     3481 cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac
     3541 agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt
     3601 tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag tgtataattc
     3661 ttgaagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     3721 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     3781 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     3841 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     3901 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     3961 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     4021 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     4081 tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg caagagcaac tcggtcgccg
     4141 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     4201 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     4261 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     4321 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     4381 aaacgacgag cgtgacacca cgatgcctgc agcaatggca acaacgttgc gcaaactatt
     4441 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     4501 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     4561 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     4621 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     4681 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     4741 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     4801 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     4861 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     4921 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     4981 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     5041 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     5101 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     5161 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     5221 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     5281 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     5341 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     5401 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     5461 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     5521 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     5581 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     5641 cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct
     5701 gtgcggtatt tcacaccgca taaattccga caccatcgaa tggtgcaaaa cctttcgcgg
     5761 tatggcatga tagcgcccgg aagagagtca attcagggtg gtgaatgtga aaccagtaac
     5821 gttatacgat gtcgcagagt atgccggtgt ctcttatcag accgtttccc gcgtggtgaa
     5881 ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct
     5941 gaattacatt cccaaccgcg tggcacaaca actggcgggc aaacagtcgt tgctgattgg
     6001 cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc
     6061 tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga
     6121 agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa
     6181 ctatccgctg gatgaccagg atgccattgc tgtggaagct gcctgcacta atgttccggc
     6241 gttatttctt gatgtctctg accagacacc catcaacagt attattttct cccatgaaga
     6301 cggtacgcga ctgggcgtgg agcatctggt cgcattgggt caccagcaaa tcgcgctgtt
     6361 agcgggccca ttaagttctg tctcggcgcg tctgcgtctg gctggctggc ataaatatct
     6421 cactcgcaat caaattcagc cgatagcgga acgggaaggc gactggagtg ccatgtccgg
     6481 ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt cccactgcga tgctggttgc
     6541 caacgatcag atggcgctgg gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg
     6601 tgcggatatc tcggtagtgg gatacgacga taccgaagac agctcatgtt atatcccgcc
     6661 gtcaaccacc atcaaacagg attttcgcct gctggggcaa accagcgtgg accgcttgct
     6721 gcaactctct cagggccagg cggtgaaggg caatcagctg ttgcccgtct cactggtgaa
     6781 aagaaaaacc accctggcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     6841 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
     6901 ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc
     6961 gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg
     7021 attacggatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     7081 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     7141 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggcg ctttgcctgg
     7201 tttccggcac cagaagcggt gccggaaagc tggctggagt gcgatcttcc tgaggccgat
     7261 actgtcgtcg tcccctcaaa ctggcagatg cacggttacg atgcgcccat ctacaccaac
     7321 gtaacctatc ccattacggt caatccgccg tttgttccca cggagaatcc gacgggttgt
     7381 tactcgctca catttaatgt tgatgaaagc tggctacagg aaggccagac gcgaattatt
     7441 tttgatggcg ttggaatt
//
