LOCUS       sequence-459925-        8713 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial expression vector of MBD2 C-terminal tagged to the 
            intein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8713)
  TITLE     MBD2 modulates heterchromatin architecture and composition
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8713)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pTYB1-MBD2 pc4786
FEATURES             Location/Qualifiers
     source          1..8713
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(782..801)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     942..964
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(958..1149)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1706..2788)
                     /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     2750..2769
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2789..2866)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      3019..3105
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3202..3288
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3385..3471
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3568..3654
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3751..3837
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3972..3991
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3972..3990
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3991..4015
                     /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             4030..4052
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5347..6165
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae VMA1"
                     /product="5' region of the intein from the yeast Vma1 
                     subunit of the vacuolar ATPase (Chong et al., 1998)"
                     /label=Sce VMA intein 5' region
                     /translation="CFAKGTNVLMADGSIECIENIEVGNKVMGKDGRPREVIKLPRGRE
                     TMYSVVQKSQHRAHKSDSSREVPELLKFTCNATHELVVRTPRSVRRLSRTIKGVEYFEV
                     ITFEMGQKKAPDGRIVELVKEVSKSYPISEGPERANELVESYRKASNKAYFEWTIEARD
                     LSLLGSHVRKATYQTYAPILYENDHFFDYMQKSKFHLTIEGPKVLAYLLGLWIGDGLSD
                     RATFSVDSRDTSLMERVTEYAEKLNLCAEYKDRKEPQVAKTVNLYSKVVRG"
     CDS             6621..6632
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             6742..6897
                     /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(6962..6980)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6976..7023
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        7083..7187
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7188..7256,7257..8048)
                     /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(7406..7425)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(8090..8603)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8285..8306)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     8497..8516
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagctatgg tgcactctca
      781 gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga
      841 ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
      901 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
      961 gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg
     1021 gtcgtgcagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc
     1081 cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg ttttttcctg
     1141 tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa tgataccgat
     1201 gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc ggttactgga
     1261 acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa aaatcactca
     1321 gggtcaatgc cagccgaacg ccagcaagac gtagcccagc gcgtcggccg ccatgccggc
     1381 gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc
     1441 gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg
     1501 aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat
     1561 gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga
     1621 gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag
     1681 ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     1741 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca
     1801 gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc
     1861 cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt
     1921 tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta
     1981 ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcacccagcg
     2041 ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca
     2101 tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa
     2161 tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac
     2221 ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc
     2281 ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga
     2341 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
     2401 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
     2461 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
     2521 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
     2581 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
     2641 tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa
     2701 cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg
     2761 cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc
     2821 gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc
     2881 tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc
     2941 accgccgccg caaggaatgg tgcatgccgg catgccgccc tttcgtcttc aagaattaat
     3001 tcccaattcc ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt
     3061 tttatctgtt gtttgtcggt gaacgctctc ctgagtagga caaatccgcc gggagcggat
     3121 ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag gacgcccgcc ataaactgcc
     3181 aggaattaat tccccaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt
     3241 cgttttatct gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc gccgggagcg
     3301 gatttgaacg ttgcgaagca acggcccgga gggtggcggg caggacgccc gccataaact
     3361 gccaggaatt aattccccag gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc
     3421 tttcgtttta tctgttgttt gtcggtgaac gctctcctga gtaggacaaa tccgccggga
     3481 gcggatttga acgttgcgaa gcaacggccc ggagggtggc gggcaggacg cccgccataa
     3541 actgccagga attaattccc caggcatcaa ataaaacgaa aggctcagtc gaaagactgg
     3601 gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc tgagtaggac aaatccgccg
     3661 ggagcggatt tgaacgttgc gaagcaacgg cccggagggt ggcgggcagg acgcccgcca
     3721 taaactgcca ggaattaatt ccccaggcat caaataaaac gaaaggctca gtcgaaagac
     3781 tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg
     3841 ccgggagcgg atttgaacgt tgcgaagcaa cggcccggag ggtggcgggc aggacgcccg
     3901 ccataaactg ccaggaattg gggatcggaa ttaattcccg gtttaaaccg gggatctcga
     3961 tcccgcgaaa ttaatacgac tcactatagg ggaattgtga gcggataaca attcccctct
     4021 agaaataatt ttgtttaact ttaagaagga gatatacata tgtcaaaggc ctacgtcgac
     4081 gccatgcgcg cgcacccggg gggaggccgc tgctgcccgg agcaggagga gggggagagc
     4141 gcggcgggcg gcagcggcgc tggcggcgac tccgccatag agcagggggg ccagggcagc
     4201 gcgctcgctc cgtccccggt gagcggcgtg cgcagggaag gcgctcgggg cggcggccgt
     4261 ggccgggggc ggtggaagca ggcggcccgg ggcggcggcg tctgtggccg tggccgtggc
     4321 cgtggccggg gtcggggccg tggccggggc cggggccggg gccgcggccg tccccagagt
     4381 ggcggcagcg gccttggcgg cgacggcggc ggcggcgcgg gcggctgcgg cgtcggcagc
     4441 ggtggcggcg tcgccccccg gcgggatcct gtccctttcc cgtcggggag ctcggggccg
     4501 gggcccaggg gaccccgggc cacggagagc gggaagagga tggactgccc ggccctcccc
     4561 cccggatgga agaaggagga agtgatccga aaatcagggc tcagtgctgg caagagcgat
     4621 gtctactact tcagtccaag tggtaagaag ttcagaagta aacctcagct ggcaagatac
     4681 ctgggaaatg ctgttgacct tagcagtttt gacttcagga ccggcaagat gatgcctagt
     4741 aaattacaga agaacaagca gagactccgg aatgaccccc tcaatcagaa caagggtaaa
     4801 ccagacctga acacaacatt gccaattaga caaactgcat caattttcaa gcaaccagta
     4861 accaaattca cgaaccaccc gagcaataag gtgaagtcag acccccagcg gatgaatgaa
     4921 caaccacgtc agcttttctg ggagaagagg ctacaaggac ttagcgcatc agatgtaaca
     4981 gaacaaatta taaaaaccat ggagctacct aaaggtcttc aaggagtcgg tccaggtagc
     5041 aatgacgaga cccttctgtc tgctgtggcc agtgctttac acacaagctc tgcgcccatc
     5101 acaggacaag tctctgctgc cgtggaaaag aaccctgctg tttggcttaa cacatctcaa
     5161 cccctctgca aagctttcat tgttacagat gaagacatta ggaaacagga agagcgagtc
     5221 caacaagtac gcaagaaact ggaggaggca ctgatggccg acatcctgtc ccgggctgcg
     5281 gacacggagg aagtagacat tgacatggac gatggagatg aggcggaatt cctcgagggc
     5341 tcttcctgct ttgccaaggg taccaatgtt ttaatggcgg atgggtctat tgaatgtatt
     5401 gaaaacattg aggttggtaa taaggtcatg ggtaaagatg gcagacctcg tgaggtaatt
     5461 aaattgccca gaggaagaga aactatgtac agcgtcgtgc agaaaagtca gcacagagcc
     5521 cacaaaagtg actcaagtcg tgaagtgcca gaattactca agtttacgtg taatgcgacc
     5581 catgagttgg ttgttagaac acctcgtagt gtccgccgtt tgtctcgtac cattaagggt
     5641 gtcgaatatt ttgaagttat tacttttgag atgggccaaa agaaagcccc cgacggtaga
     5701 attgttgagc ttgtcaagga agtttcaaag agctacccaa tatctgaggg gcctgagaga
     5761 gccaacgaat tagtagaatc ctatagaaag gcttcaaata aagcttattt tgagtggact
     5821 attgaggcca gagatctttc tctgttgggt tcccatgttc gtaaagctac ctaccagact
     5881 tacgctccaa ttctttatga gaatgaccac tttttcgact acatgcaaaa aagtaagttt
     5941 catctcacca ttgaaggtcc aaaagtactt gcttatttac ttggtttatg gattggtgat
     6001 ggattgtctg acagggcaac tttttcggtt gattccagag atacttcttt gatggaacgt
     6061 gttactgaat atgctgaaaa gttgaatttg tgcgccgagt ataaggacag aaaagaacca
     6121 caagttgcca aaactgttaa tttgtactct aaagttgtca gaggtaatgg tattcgcaat
     6181 aatcttaata ctgagaatcc attatgggac gctattgttg gcttaggatt cttgaaggac
     6241 ggtgtcaaaa atattccttc tttcttgtct acggacaata tcggtactcg tgaaacattt
     6301 cttgctggtc taattgattc tgatggctat gttactgatg agcatggtat taaagcaaca
     6361 ataaagacaa ttcatacttc tgtcagagat ggtttggttt cccttgctcg ttctttaggc
     6421 ttagtagtct cggttaacgc agaacctgct aaggttgaca tgaatgtcac caaacataaa
     6481 attagttatg ctatttatat gtctggtgga gatgttttgc ttaacgttct ttcgaagtgt
     6541 gccggctcta aaaaattcag gcctgctccc gccgctgctt ttgcacgtga gtgccgcgga
     6601 ttttatttcg agttacaaga attgaaggaa gacgattatt atgggattac tttatctgat
     6661 gattctgatc atcagttttt gcttggatcc caggttgtcg tccatgcatg cggtggcctg
     6721 accggtctga actcaggcct cacgacaaat cctggtgtat ccgcttggca ggtcaacaca
     6781 gcttatactg cgggacaatt ggtcacatat aacggcaaga cgtataaatg tttgcagccc
     6841 cacacctcct tggcaggatg ggaaccatcc aacgttcctg ccttgtggca gcttcaatga
     6901 ctgcaggaag gggatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc
     6961 accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt
     7021 ttgctgaaag gaggaactat atccggataa ctacgtcagg tggcactttt cggggaaatg
     7081 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     7141 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     7201 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     7261 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     7321 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgtttcc
     7381 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt gttgacgccg
     7441 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     7501 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     7561 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     7621 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     7681 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     7741 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     7801 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     7861 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     7921 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     7981 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     8041 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta ccccggttga
     8101 taatcagaaa agccccaaaa acaggaagat tgtataagca aatatttaaa ttgtaaacgt
     8161 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     8221 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag cccgagatag ggttgagtgt
     8281 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     8341 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcacccaaat caagtttttt
     8401 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     8461 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     8521 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     8581 taatgcgccg ctacagggcg cgtaaaagga tctaggtgaa gatccttttt gataatctca
     8641 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     8701 tcaaaggatc ttc
//
