LOCUS Exported 7306 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Expresses GST-EndoE(E662Q) from E. faecalis in E. coli. ACCESSION . VERSION . KEYWORDS pGEXndoE(E662Q) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7306) AUTHORS Collin M, Fischetti VA TITLE A novel secreted endoglycosidase from Enterococcus faecalis with activity on human immunoglobulin G and ribonuclease B. JOURNAL J Biol Chem. 2004 May 21;279(21):22558-70. Epub 2004 Mar 17. PUBMED 15028731 REFERENCE 2 (bases 1 to 7306) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7306 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 183..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)." CDS 258..911 /codon_start=1 /product="glutathione S-transferase from Schistosoma japonicum" /label=GST /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK" primer_bind 869..891 /label=pGEX 5' /note="pGEX vectors, Glutathione-S-transferase, forward primer" CDS 921..932 /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(3356..3378) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(3523..3541) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 3609..3713 /gene="bla" /label=AmpR promoter CDS 3714..4574 /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(3932..3951) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4745..5333 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5234..5253 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5487..5504 /label=L4440 /note="L4440 vector, forward primer" promoter 5577..5654 /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 5655..6737 /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(5674..5693) /label=LacI-R /note="LacI, reverse primer" protein_bind 6750..6771 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 6786..6816 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6824..6840 /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 6848..6864 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 6848..6864 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" CDS 6860..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(6880..6897) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(6880..6896) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(6889..6911) /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 atcctccaaa atcggatctg atcgaaggtc gtgggatccc caatgagccg acccaagaga 961 aacattttat ggtttattat cgggcttggc gtgacaaaac gatgcaagga gttaatacaa 1021 cattgccaga tgaaaattgg ctaacgatgc acgatattcc ttatggtatt gacattgtga 1081 atgtctttag ttatgtgcca aaaggacaag aagcacttgc acagccattt tatgatacgt 1141 taaaaaatga gtatgcgcca gcactgcatg cacgaggtgt tcgtttagtt cgagggattg 1201 attacagcga gctattaaaa gttccttatg caggaacaac gcctacagaa gcagaatttg 1261 atgcttatgc gaaagagttg ttaaccaaat ttgtcgatga tttaggaatt gatgggttag 1321 atattgacat ggaaactcgt ccaagtgaaa aagatattgt gctatctaat ggtgtcattc 1381 gtgcattatc aaaatacatt ggaccgaagt cgggaacgga tcgtccattt ttgtatgata 1441 ccaatgcaga atatttacca cctttacaag atgtcagtga ctgtttcgat tttctcgcgt 1501 atcaacagta tggtagcgat gacaaacgca cgcaacgagc attaaataat ttaagtccgg 1561 ttctcaatgg ggaacgattt gttccaggat taactttccc agaagagcaa gatcgcaacc 1621 gctggtatga cacaaaagag ccgtatatgg aaagtaacat gtataaagta gctcgttatt 1681 cttatgaaaa taatttaggg ggcatgttcc tctatgcctt agatcgcgat ggtcgcacct 1741 ataatgaaga cgatttaaat cagataaaac cttctaattt actttggaca aaaacggcca 1801 ttgcagagag taaaggcgtt tctcttgcag agatgaaagc ggctgcgcaa cactatttaa 1861 aacgaattag ctacgccaac acagaccttg aagcacaaaa taaagccgca gaagcagtga 1921 cacaagcaac aacgctttat gatgtgaata aagctatttt aggtggcgat tatggccaag 1981 ggctttcaaa tacctatgat gctgaattag aaaaaggcct gttagccatt gacttaacca 2041 ctttatatcg tgcgttggat caagcagttg cagccattga aaaggcagaa agctatacac 2101 cagaaacgat tcaagcacta caaacaacaa aagagtcagt cgcaacagaa cttgcgggaa 2161 aaacgtatac agccgcgcaa gtgactactt ggcaaacgga ggtccaaaca gctttggata 2221 atttaaaaga gaaacaaaca caacctttaa aaagtgtctt ctccattgat gcgggaagaa 2281 aatatttttc cgtggaacaa ctggaagaat tagtggccaa agccagtcaa aatgggtaca 2341 cagatgtcca attaatttta ggaaatgatg gcttacggtt tattttggat gatatgtcgg 2401 tcaatgtgaa tggtaaaaaa tacaaccaca accgggtttc aaaagcaatc caacgaggca 2461 acaacgcata ttacaatgat cctaatggca acgcgttaac acaaaaagaa atggatcggt 2521 tgttggcttt tgcgaaagcg cgcaacatca acatcattcc cgtgattaac agtccaggac 2581 atatggatgc gttgttagtg gccatggaaa aactagcgat taaaaatccc gcttttgatg 2641 gctcaaaacg aacagtagat ttagggaacc aaaaggcagt gaatttcaca aaggcgatta 2701 tcagtaagta cgtggcttat ttttccgcgc atagtgaaat tttcaatttt ggtggcgatc 2761 agtatgcaaa tgatgtcgac acaggcggtt gggcgaaact gcaatcttct gggcgctaca 2821 aggattttgt cgcttatgcg aatgatttag ctaaaataat taaagatgcg ggcatgcagc 2881 caatgagctt taatgatggc atttattaca acagcgacga ttctttcggt acatttgacc 2941 cagagattat tatttcttat tggacagccg gttggagtgg gtatgacgta gccaaacctg 3001 agtactttgt tcaaaaagga cacaaaattt ttaataccaa tgatgcgtgg tattgggtcg 3061 ctggcaatgt tgattctggc atttatcaat atgacgatgc tttagcaaat atgtcgaaaa 3121 aagcatttac agatgtgcca gcaggtagcc caaatcttcc aattattgga agtattcaat 3181 gtgtttggta tgatgaccct cgtcgtgact atgattttga acgaatttat acgctaatgg 3241 atacgttctc ggaaaattat cgtgagtata tggtggttaa aaatcactag ctcgagcggc 3301 cgcatcgtga ctgactgacg atctgcctcg cgcgtttcgg tgatgacggt gaaaacctct 3361 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac 3421 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt 3481 cacgtagcga tagcggagtg tataattctt gaagacgaaa gggcctcgtg atacgcctat 3541 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg 3601 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc 3661 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta 3721 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg 3781 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg 3841 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac 3901 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtgttg 3961 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt 4021 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg 4081 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac 4141 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt 4201 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag 4261 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc 4321 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc 4381 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta 4441 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg 4501 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga 4561 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac 4621 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa 4681 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat 4741 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc 4801 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg 4861 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc 4921 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg 4981 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg 5041 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa 5101 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg 5161 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga 5221 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct 5281 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca 5341 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc 5401 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg 5461 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc 5521 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata aattccgaca 5581 ccatcgaatg gtgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat 5641 tcagggtggt gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct 5701 cttatcagac cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg 5761 aaaaagtgga agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac 5821 tggcgggcaa acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc 5881 cgtcgcaaat tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg 5941 tgtcgatggt agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg 6001 cgcaacgcgt cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg 6061 tggaagctgc ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca 6121 tcaacagtat tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg 6181 cattgggtca ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc 6241 tgcgtctggc tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac 6301 gggaaggcga ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg 6361 gcatcgttcc cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg 6421 ccattaccga gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata 6481 ccgaagacag ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc 6541 tggggcaaac cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca 6601 atcagctgtt gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa 6661 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac 6721 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc 6781 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa 6841 tttcacacag gaaacagcta tgaccatgat tacggattca ctggccgtcg ttttacaacg 6901 tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt 6961 cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag 7021 cctgaatggc gaatggcgct ttgcctggtt tccggcacca gaagcggtgc cggaaagctg 7081 gctggagtgc gatcttcctg aggccgatac tgtcgtcgtc ccctcaaact ggcagatgca 7141 cggttacgat gcgcccatct acaccaacgt aacctatccc attacggtca atccgccgtt 7201 tgttcccacg gagaatccga cgggttgtta ctcgctcaca tttaatgttg atgaaagctg 7261 gctacaggaa ggccagacgc gaattatttt tgatggcgtt ggaatt //