LOCUS       sequence-479129-        8726 bp    DNA     circular SYN 22-JUL-2025
DEFINITION  Protein expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8726)
  AUTHORS   Goulas T, Cuppari A, Garcia-Castellanos R, Snipas S, Glockshuber R, 
            Arolas JL, Gomis-Ruth FX
  TITLE     The pCri System: A Vector Collection for Recombinant Protein 
            Expression and Purification.
  JOURNAL   PLoS One. 2014 Nov 11;9(11):e112643. doi: 
            10.1371/journal.pone.0112643. eCollection 2014.
  PUBMED    25386923
REFERENCE   2  (bases 1 to 8726)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 22, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS One.";
            date: "2014-11-11"; volume: "9(11):e112643. doi"; pages: " 
            10.1371/journal.pone.0112643. eCollection 2014"
COMMENT     Sequence Label: pCri-17a
FEATURES             Location/Qualifiers
     source          1..8726
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..25
                     /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             59..76
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             86..106
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(119..121,122..835)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIHHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(161..182)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     769..790
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             887..904
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        4043..4147
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4148..4216,4217..5008)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4366..4385)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5179..5767
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5668..5687
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5921..5938
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7187..7209
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7201..7218
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7202..7218
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(7261..8343)
                     /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
                     EACKTAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     8305..8324
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ggaattgtga gcggataaca attcccaatt aaaggaggaa gtcaagatgg gcagcagcca
       61 tcatcatcat catcacagca gcggcgaaaa cctttacttc cagggcccca tggccaaagt
      121 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
      181 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
      241 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
      301 gaccaccttc ggctacggcc tgcagtgctt cgcccgctac cccgaccaca tgaagcagca
      361 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
      421 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
      481 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
      541 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
      601 caaggtgaac ttcaagatcc accacaacat cgaggacggc agcgtgcagc tcgccgacca
      661 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
      721 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
      781 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtaaggtac
      841 cggatccgaa ttcgagctcc gtcgacaagc ttgcggccca ctcgagcacc accaccacca
      901 ccactgaaaa gtttatacct tacctggaac aaatggttga aacatacgag gctaatatcg
      961 gcttattagg aatagtccct gtactaataa aatcaggtgg atcagttgat cagtatattt
     1021 tggacgaagc tcggaaagaa tttggagatg acttgcttaa ttccacaatt aaattaaggg
     1081 aaagaataaa gcgatttgat gttcaaggaa tcacggaaga agatactcat gataaagaag
     1141 ctctaaaact attcaataac cttacaatgg aattgatcga aagggtggaa ggttaatggt
     1201 acgaaaatta ggggatctac ctagaaagcc acaaggcgat aggtcaagct taaagaaccc
     1261 ttacatggat cttacagatt ctgaaagtaa agaaacaaca gaggttaaac aaacagaacc
     1321 aaaaagaaaa aaagcattgt tgaaaacaat gaaagttgat gtttcaatcc ataataagat
     1381 taaatcgctg cacgaaattc tggcagcatc cgaagggaat tcatattact tagaggatac
     1441 tattgagaga gctattgata agatggttga gacattacct gagagccaaa aaacttttta
     1501 tgaatatgaa ttaaaaaaaa gaaccaacaa aggctgagac agactccaaa cgagtctgtt
     1561 tttttaaaaa aaatattagg agcattgaat atatattaga gaattaagaa agacatggga
     1621 ataaaaatat tttaaatcca gtaaaaatat gataagatta tttcagaata tgaagaactc
     1681 tgtttgtttt tgatgaaaaa acaaacaaaa aaaatccacc taacggaatc tcaatttaac
     1741 taacagcggc caaactgaga agttaaattt gagaagggga aaaggcggat ttatacttgt
     1801 atttaactat ctccatttta acattttatt aaaccccata caagtgaaaa tcctctttta
     1861 cactgttcct ttaggtgatc gcggagggac attatgagtg aagtaaacct aaaaggaaat
     1921 acagatgaat tagtgtatta tcgacagcaa accactggaa ataaaatcgc caggaagaga
     1981 atcaaaaaag ggaaagaaga agtttattat gttgctgaaa cggaagagaa gatatggaca
     2041 gaagagcaaa taaaaaactt ttctttagac aaatttggta cgcatatacc ttacatagaa
     2101 ggtcattata caatcttaaa taattacttc tttgattttt ggggctattt tttaggtgct
     2161 gaaggaattg cgctctatgc tcacctaact cgttatgcat acggcagcaa agacttttgc
     2221 tttcctagtc tacaaacaat cgctaaaaaa atggacaaga ctcctgttac agttagaggc
     2281 tacttgaaac tgcttgaaag gtacggtttt atttggaagg taaacgtccg taataaaacc
     2341 aaggataaca cagaggaatc cccgattttt aagattagac gtaaggttcc tttgctttca
     2401 gaagaacttt taaatggaaa ccctaatatt gaaattccag atgacgagga agcacatgta
     2461 aagaaggctt taaaaaagga aaaagagggt cttccaaagg ttttgaaaaa agagcacgat
     2521 gaatttgtta aaaaaatgat ggatgagtca gaaacaatta atattccaga ggccttacaa
     2581 tatgacacaa tgtatgaaga tatactcagt aaaggagaaa ttcgaaaaga aatcaaaaaa
     2641 caaataccta atcctacaac atcttttgag agtatatcaa tgacaactga agaggaaaaa
     2701 gtcgacagta ctttaaaaag cgaaatgcaa aatcgtgtct ctaagccttc ttttgatacc
     2761 tggtttaaaa acactaagat caaaattgaa aataaaaatt gtttattact tgtaccgagt
     2821 gaatttgcat ttgaatggat taagaaaaga tatttagaaa caattaaaac agtccttgaa
     2881 gaagctggat atgttttcga aaaaatcgaa ctaagaaaag tgcaataaac tgctgaagta
     2941 tttcagcagt tttttttatt tagaaatagt gaaaaaaata taatcaggga ggtatcaata
     3001 tttaatgagt actgatttaa atttatttag actggaatta ataattaaca cgtagactaa
     3061 ttaaaattta atgagggata aagaggatac aaaaatatta atttcaatcc ctattaaatt
     3121 ttaacaaggg ggggattaaa atttaattag aggtttatcc acaagaaaag accctaataa
     3181 aatttttact agggttataa cactgattaa tttcttaatg ggggagggat taaaatttaa
     3241 tgacaaagaa aacaatcttt taagaaaagc ttttaaaaga taataataaa aagagctttg
     3301 cgattaagca aaactcttta ctttttcatt gacattatca aattcatcga tttcaaattg
     3361 ttgttgtatc ataaagttaa ttctgttttg cacaaccttt tcaggaatat aaaacacatc
     3421 tgaggcttgt tttataaact cagggtcgct aaagtcaatg taacgtagca tatgatatgg
     3481 tatagcttcc acccaagtta gcctttctgc ttcttctgaa tgtttttcat atacttccat
     3541 cggtatctct aaatgatttt cctcatgtag caaggtatga gcaaaaagtt tatggaattg
     3601 atagttcctc tctttttctt caactttttt atctaaaaca aacactttaa catctgagtc
     3661 aatgtaagca taagatgttt ttccagtcat aatttcaatc ccaaatcttt tagacagaaa
     3721 ttctggacgt aaatcttttg gtgaaagaat ttttttatgt agcaatatat ccgatacagc
     3781 accttctaaa agcgttggtg aatagggcat tttacctatc tcctctcatt ttgtggaata
     3841 aaaatagtca tattcgtcca tctacctatc ctattatcga acagttgaac tttttaatca
     3901 aggatcagtc ctttttttca ttattcttaa actgtgctct taactttaac aactcgattt
     3961 gtttttccag attcgagggt aactagcctc gccgatcccg caagaggccc ggcagtcagg
     4021 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     4081 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     4141 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     4201 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     4261 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     4321 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     4381 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     4441 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     4501 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     4561 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     4621 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     4681 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     4741 tctagcttcc cggcaacaat taatagactg gatggaggca gataaagttg caggaccact
     4801 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     4861 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     4921 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     4981 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     5041 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     5101 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     5161 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     5221 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     5281 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     5341 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     5401 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     5461 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     5521 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     5581 cgccacgctt cccaaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     5641 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     5701 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     5761 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     5821 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     5881 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     5941 agcggaagag cgcccaatac gcatgcttaa gttattggta tgactggttt taagcgcaaa
     6001 aaaagttgct ttttcgtacc tattaatgta tcgttagaaa accgactgta aaaagtacag
     6061 tcggcattat ctcatattat aaaagccagt cattaggcct atctgacaat tcctgaatag
     6121 agttcataaa caatcctgca tgataaccat cacaaacaga atgatgtacc tgtaaagata
     6181 gcggtaaata tattgaatta cctttattaa tgaattttcc tgctgtaata atgggtagaa
     6241 ggtaattact attattattg atatttaagt taaacccagt aaatgaagtc caaggaataa
     6301 tagaaagaga aaaagcattt tcaggtatag gtgttttggg aaacaatttc cccgaaccat
     6361 tatatttctc tacatcagaa aggtataaat cataaaactc tttgaagtca ttctttacag
     6421 gagtccaaat accagagaat gttttagata caccatcaaa aattgtataa agtggctcta
     6481 acttatccca ataacctaac tctccgtcgc tattgtaacc agttctaaaa gctgtatttg
     6541 agtttatcac ccttgtcact aagaaaataa atgcagggta aaatttatat ccttcttgtt
     6601 ttatgtttcg gtataaaaca ctaatatcaa tttctgtggt tatactaaaa gtcgtttgtt
     6661 ggttcaaata atgattaaat atctcttttc tcttccaatt gtctaaatca attttattaa
     6721 agttcatttg atatgcctcc taaattttta tctaaagtga atttaggagg cttacttgtc
     6781 tgctttcttc attagaatca atcctttttt aaaagtcaat attactgtaa cataaatata
     6841 tattttaaaa atatcccact ttatccaatt tttgtttgtt gaactaatgg gtgctttagt
     6901 tgaagaataa aagaccacat taaaaaatgt ggtcttttgt gtttttttaa aggatttgag
     6961 cgtagcgaaa aatccttttc tttcttatct tgataataag ggtaactatt gccgtcgtcc
     7021 attccgacag catcgccagt cactatggcg tgctgctagc gccattcgcc attcaggctg
     7081 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa
     7141 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt
     7201 tgtaaaacga cggccagtga attcgagctc aggccttaac tcacattaat tgcgttgcgc
     7261 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     7321 cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga
     7381 gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc
     7441 cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata
     7501 acatgagctg tcttcggtat cgtcgtatcc cactaccgag atatccgcac caacgcgcag
     7561 cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat
     7621 cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc
     7681 actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg
     7741 ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat
     7801 ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga
     7861 gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt
     7921 agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag
     7981 cccactgacg cgttgcgcga gaagattgtg caccgccgtt ttacaggctt cgacgccgct
     8041 tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc
     8101 cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa
     8161 cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat
     8221 cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg
     8281 ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt
     8341 catcaaaatc gtctccctcc gtttgaatat ttgattgatc gtaaccagat gaagcactct
     8401 ttccactatc cctacagtgt tatggcttga acaatcacga aacaataatt ggtacgtacg
     8461 atctttcagc cgactcaaac atcaaatctt acaaatgtag tctttgaaag tattacatat
     8521 gtaagattta aatgcaaccg ttttttcgga aggaaatgat gacctcgttt ccaccggaat
     8581 tagcttggta ccagctattg taacataatc ggtacggggg tgaaaaagct aacggaaaag
     8641 ggagcggaaa agaatgatgt aagcgtgaaa aattttttat cttatcactt gaaattggaa
     8701 gggagattct ttattataag aattgt
//
