LOCUS       sequence-311794-        6331 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Bacterial Expression of 6xHis HOXD13 (+10A) IDR mCherry Fusion 
            Protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6331)
  AUTHORS   Basu S, Mackowiak SD, Niskanen H, Knezevic D, Asimi V, Grosswendt S,
            Geertsema H, Ali S, Jerkovic I, Ewers H, Mundlos S, Meissner A, 
            Ibrahim DM, Hnisz D
  TITLE     Unblending of Transcriptional Condensates in Human Repeat Expansion 
            Disease.
  JOURNAL   Cell. 2020 May 28;181(5):1062-1079.e30. doi: 
            10.1016/j.cell.2020.04.018. Epub 2020 May 7.
  PUBMED    32386547
REFERENCE   2  (bases 1 to 6331)
  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: 
            "10.1016/j.cell.2020.04.018"; journalName: "Cell"; date: 
            "2020-05-28- 28"; volume: "181"; issue: "5"; pages: "1062-1079.e30"
COMMENT     Sequence Label: pET-45b-HOXD13(+10A)-IDR-mCherry
FEATURES             Location/Qualifiers
     source          1..6331
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             1..23
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             36..53
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1239..1283
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1321..1339)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1335..1382
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1419..1874
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1506..1525)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1716..1737
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(1989..2780,2781..2849))
                     /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     2612..2631
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2850..2942)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3023..3611
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3512..3531
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3765..3782
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3797..3936
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3862..3881)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4022..4044
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4038..4229)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4801..5883)
                     /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     5845..5864
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5884..5961)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6167..6186
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6274..6293
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6274..6292
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6293..6317
                     /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)."
ORIGIN
        1 tttgtttaac tttaagaagg agatatacca tggcacatca ccaccaccat cacgtgggta
       61 ccggtgacag tggatccgga gctaccatgt cccgagccgg tagctgggac atggacggcc
      121 tgcgagcgga cggaggcgga gctggaggag ctcccgcatc aagtagtagc tcaagtgtgg
      181 ccgctgccgc ggctagcgga caatgtaggg ggtttttgtc agcgcctgtc tttgcgggca
      241 cacattccgg gagggccgct gctgcggctg ctgctgctgc tgcagccgcg gcggccgcag
      301 ctgcagctgc ggccgcagca gctgccgcta gtggatttgc ctatcctggg acgagtgagc
      361 gcactggttc atcatcctca tcttcatcca gtgcggtagt cgccgctcgg ccagaagcac
      421 cacctgcaaa agagtgtccg gccccgacgc cagctgcagc agcagcggca ccgccttcag
      481 ctcctgcggg taccggaggc gggatggttt ccaaaggaga ggaggataat atggctataa
      541 ttaaagagtt tatgcggttc aaggtgcata tggagggatc tgtgaacggt cacgagttcg
      601 aaatcgaggg cgaaggcgag gggcgaccct atgaagggac acagacggcc aaactgaagg
      661 tgaccaaggg tggccccctg cctttcgcct gggacatcct gtcaccccag tttatgtacg
      721 gctctaaggc ttacgttaag caccctgccg atattccgga ctacttgaaa ctgagcttcc
      781 ctgaagggtt caagtgggaa cgggtaatga attttgagga tggcggtgta gtcacagtta
      841 cccaggacag ctctcttcag gacggagaat ttatctataa ggttaaactt cggggcacta
      901 acttcccatc agacggcccc gtgatgcaga agaaaactat gggctgggaa gccagctcag
      961 aacgcatgta tcccgaggac ggagcactga agggagaaat caagcagcga ctcaagctga
     1021 aggatggggg acattatgat gctgaggtca aaaccaccta taaggccaag aaacccgttc
     1081 agcttcctgg ggcatacaac gtgaatatca aactggatat tacatcccat aacgaagact
     1141 ataccatcgt ggaacagtac gagcgggccg agggcagaca tagcacaggg ggcatggatg
     1201 aattgtacaa gaagcttgcg gccgcactcg agtctggtaa agaaaccgct gctgcgaaat
     1261 ttgaacgcca gcacatggac tcgtctacta gcgcagctta attaacctag gctgctgcca
     1321 ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     1381 tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta gcggcgcatt
     1441 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     1501 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     1561 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     1621 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     1681 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     1741 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     1801 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     1861 aacgtttaca atttctggcg gcacgatggc atgagattat caaaaaggat cttcacctag
     1921 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     1981 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     2041 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     2101 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     2161 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     2221 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     2281 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     2341 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     2401 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     2461 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     2521 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     2581 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     2641 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     2701 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     2761 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     2821 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatcatg attgaagcat
     2881 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     2941 aataggtcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     3001 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     3061 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     3121 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     3181 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     3241 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     3301 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     3361 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     3421 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     3481 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     3541 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     3601 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     3661 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     3721 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     3781 aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3841 accgcaatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac
     3901 actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct
     3961 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     4021 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg
     4081 cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg
     4141 tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg
     4201 ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc
     4261 atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat
     4321 gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg
     4381 gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt
     4441 ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct
     4501 gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct
     4561 caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca
     4621 ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg
     4681 atcatgctag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag
     4741 ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc
     4801 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     4861 cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga
     4921 gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc
     4981 cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata
     5041 acatgagctg tcttcggtat cgtcgtatcc cactaccgag atgtccgcac caacgcgcag
     5101 cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat
     5161 cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc
     5221 actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg
     5281 ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat
     5341 ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga
     5401 gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt
     5461 agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag
     5521 cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct
     5581 tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc
     5641 cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa
     5701 cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat
     5761 cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg
     5821 ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt
     5881 cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt
     5941 tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac tcctgcatta
     6001 ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat
     6061 gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga
     6121 aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga
     6181 tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt
     6241 agaggatcga gatcgatctc gatcccgcga aattaatacg actcactata ggggaattgt
     6301 gagcggataa caattcccct ctagaaataa t
//
