LOCUS       sequence-226720-        6998 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Bacterial expression plasmid encoding X. laevis H2B residues 1-114 
            as a chimeric fusion to GyrA intein with chitin binding domain at 
            the C-terminus. H2B fragment can be derivatized using MESNa.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6998)
  AUTHORS   Morgan MT, Haj-Yahya M, Ringel AE, Bandi P, Brik A, Wolberger C
  TITLE     Structural basis for histone H2B deubiquitination by the SAGA DUB 
            module.
  JOURNAL   Science. 2016 Feb 12;351(6274):725-8. doi: 10.1126/science.aac5681.
  PUBMED    26912860
REFERENCE   2  (bases 1 to 6998)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1126/science"; 
            journalName: "Science"; date: "2016-02-12- 12"; volume: "351"; 
            issue: "6274"; pages: "725-8"
COMMENT     Sequence Label: pTXB1-H2B(1-114)_wt
FEATURES             Location/Qualifiers
     source          1..6998
                     /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)."
     RBS             40..62
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             413..1006
                     /codon_start=1
                     /gene="Mycobacterium xenopi gyrA"
                     /product="modified GyrA intein (Southworth et al., 1999)"
                     /label=Mxe GyrA intein
                     /note="N-terminal cleavage of this intein can be induced by
                     thiols."
                     /translation="CITGDALVALPEGESVRIADIVPGARPNSDNAIDLKVLDRHGNPV
                     LADRLFHSGEHPVYTVRTVEGLRVTGTANHPLLCLVDVAGVPTLLWKLIDEIKPGDYAV
                     IQRSAFSVDCAGFARGKPEFAPTTYTVGVPGLVRFLEAHHRDPDAQAIADELTDGRFYY
                     AKVASVTDAGVQPVYSLRVDTADHAFITNGFVSHA"
     CDS             1037..1192
                     /codon_start=1
                     /gene="Bacillus circulans chiA"
                     /product="chitin binding domain from chitinase A1 (Watanabe
                     et al., 1994)"
                     /label=CBD
                     /translation="TTNPGVSAWQVNTAYTAGQLVTYNGKTYKCLQPHTSLAGWEPSNV
                     PALWQLQ"
     primer_bind     complement(1257..1275)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1271..1318
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        1378..1482
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1483..1551,1552..2343)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1701..1720)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(2385..2898)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2580..2601)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2792..2811
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      3009..3597
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3498..3517
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3751..3768
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3790..3809)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3950..3972
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3966..4157)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4714..5796)
                     /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     5758..5777
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5797..5874)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      6027..6113
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6210..6296
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6393..6479
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6576..6662
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6759..6845
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(6980..6998,1)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6980..6998
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga
       61 gatatacata atggccaagt ccgctccagc cccgaagaaa ggctccaaga aagcggtgac
      121 caagactcag aagaaagacg ggaaaaagcg caggaagaca aggaaggaga gttatgccat
      181 ttacgtgtac aaggtgctga agcaggtgca ccccgatacc ggcatctcgt ccaaggccat
      241 gagcatcatg aactcctttg tcaacgatgt gtttgagcgc atcgcagggg aagcctcccg
      301 cctggctcat tacaacaagc gctccaccat cacctcccgg gagatccaga ccgcggtccg
      361 actgctgctg cctggggagt tggccaaaca cgccgtgtcc gagggcacca agtgcatcac
      421 gggagatgca ctagttgccc tacccgaggg cgagtcggta cgcatcgccg acatcgtgcc
      481 gggtgcgcgg cccaacagtg acaacgccat cgacctgaaa gtccttgacc ggcatggcaa
      541 tcccgtgctc gccgaccggc tgttccactc cggcgagcat ccggtgtaca cggtgcgtac
      601 ggtcgaaggt ctgcgtgtga cgggcaccgc gaaccacccg ttgttgtgtt tggtcgacgt
      661 cgccggggtg ccgaccctgc tgtggaagct gatcgacgaa atcaagccgg gcgattacgc
      721 ggtgattcaa cgcagcgcat tcagcgtcga ctgtgcaggt tttgcccgcg ggaaacccga
      781 atttgcgccc acaacctaca cagtcggcgt ccctggactg gtgcgtttct tggaagcaca
      841 ccaccgagac ccggacgccc aagctatcgc cgacgagctg accgacgggc ggttctacta
      901 cgcgaaagtc gccagtgtca ccgacgccgg cgtgcagccg gtgtatagcc ttcgtgtcga
      961 cacggcagac cacgcgttta tcacgaacgg gttcgtcagc cacgctactg gcctcaccgg
     1021 tctgaactca ggcctcacga caaatcctgg tgtatccgct tggcaggtca acacagctta
     1081 tactgcggga caattggtca catataacgg caagacgtat aaatgtttgc agccccacac
     1141 ctccttggca ggatgggaac catccaacgt tcctgccttg tggcagcttc aatgactgca
     1201 ggaaggggat ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc
     1261 tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct
     1321 gaaaggagga actatatccg gataactacg tcaggtggca cttttcgggg aaatgtgcgc
     1381 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     1441 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     1501 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     1561 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     1621 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg tttcccaatg
     1681 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtgttga cgccgggcaa
     1741 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     1801 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     1861 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     1921 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     1981 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     2041 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     2101 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     2161 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     2221 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     2281 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     2341 taactgtcag accaagttta ctcatatata ctttagattg atttaccccg gttgataatc
     2401 agaaaagccc caaaaacagg aagattgtat aagcaaatat ttaaattgta aacgttaata
     2461 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     2521 aaatcggcaa aatcccttat aaatcaaaag aatagcccga gatagggttg agtgttgttc
     2581 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     2641 ccgtctatca gggcgatggc ccactacgtg aaccatcacc caaatcaagt tttttggggt
     2701 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     2761 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     2821 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     2881 cgccgctaca gggcgcgtaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     2941 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     3001 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     3061 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     3121 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     3181 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     3241 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     3301 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     3361 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     3421 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     3481 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     3541 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     3601 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     3661 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     3721 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agctatggtg
     3781 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg
     3841 ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga
     3901 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     3961 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca
     4021 tcagcgtggt cgtgcagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg
     4081 agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt
     4141 ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg
     4201 ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga acatgcccgg
     4261 ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga ccagagaaaa
     4321 atcactcagg gtcaatgcca gccgaacgcc agcaagacgt agcccagcgc gtcggccgcc
     4381 atgccggcga taatggcctg cttctcgccg aaacgtttgg tggcgggacc agtgacgaag
     4441 gcttgagcga gggcgtgcaa gattccgaat accgcaagcg acaggccgat catcgtcgcg
     4501 ctccagcgaa agcggtcctc gccgaaaatg acccagagcg ctgccggcac ctgtcctacg
     4561 agttgcatga taaagaagac agtcataagt gcggcgacga tagtcatgcc ccgcgcccac
     4621 cggaaggagc tgactgggtt gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa
     4681 tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     4741 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     4801 gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac
     4861 cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa
     4921 atcctgtttg atggtggtta acggcgggat ataacatgag ctgtcttcgg tatcgtcgta
     4981 tcccactacc gagatatccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc
     5041 gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag
     5101 catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat
     5161 cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga
     5221 gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg
     5281 ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg
     5341 gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc
     5401 atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt
     5461 gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct
     5521 ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag
     5581 ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc
     5641 cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt
     5701 cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc
     5761 atactctgcg acatcgtata acgttactgg tttcacattc accaccctga attgactctc
     5821 ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat
     5881 ctcgacgctc tcccttatgc gactcctgca ttaggaagca gcccagtagt aggttgaggc
     5941 cgttgagcac cgccgccgca aggaatggtg catgccggca tgccgccctt tcgtcttcaa
     6001 gaattaattc ccaattcccc aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg
     6061 cctttcgttt tatctgttgt ttgtcggtga acgctctcct gagtaggaca aatccgccgg
     6121 gagcggattt gaacgttgcg aagcaacggc ccggagggtg gcgggcagga cgcccgccat
     6181 aaactgccag gaattaattc cccaggcatc aaataaaacg aaaggctcag tcgaaagact
     6241 gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc
     6301 cgggagcgga tttgaacgtt gcgaagcaac ggcccggagg gtggcgggca ggacgcccgc
     6361 cataaactgc caggaattaa ttccccaggc atcaaataaa acgaaaggct cagtcgaaag
     6421 actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc
     6481 cgccgggagc ggatttgaac gttgcgaagc aacggcccgg agggtggcgg gcaggacgcc
     6541 cgccataaac tgccaggaat taattcccca ggcatcaaat aaaacgaaag gctcagtcga
     6601 aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa
     6661 atccgccggg agcggatttg aacgttgcga agcaacggcc cggagggtgg cgggcaggac
     6721 gcccgccata aactgccagg aattaattcc ccaggcatca aataaaacga aaggctcagt
     6781 cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc ctgagtagga
     6841 caaatccgcc gggagcggat ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag
     6901 gacgcccgcc ataaactgcc aggaattggg gatcggaatt aattcccggt ttaaaccggg
     6961 gatctcgatc ccgcgaaatt aatacgactc actatagg
//
