LOCUS       sequence-272523-        6124 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lin28-His6 was overexpressed in Escherichia coli Rosetta (DE3) 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6124)
  AUTHORS   O'Day E, Le MT, Imai S, Tan SM, Kirchner R, Arthanari H, Hofmann O, 
            Wagner G, Lieberman J
  TITLE     An RNA-binding Protein, Lin28, Recognizes and Remodels G-quartets in
            the MicroRNAs (miRNAs) and mRNAs It Regulates.
  JOURNAL   J Biol Chem. 2015 Jul 17;290(29):17909-22. doi: 
            10.1074/jbc.M115.665521. Epub 2015 Jun 4.
  PUBMED    26045559
REFERENCE   2  (bases 1 to 6124)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1074/jbc.M115.665521"; journalName: "J Biol Chem"; date: 
            "2015-07-17- 17"; volume: "290"; issue: "29"; pages: "17909-22"
COMMENT     Sequence Label: pET15b-mLin28A-cHIS
FEATURES             Location/Qualifiers
     source          1..6124
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             550..567
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(626..644)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      640..687
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(862..867,868..884,885..890))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(917..935)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1003..1107
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1108..1176,1177..1968)
                     /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(1326..1345)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2139..2727
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2628..2647
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2881..2898
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2913..3052
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2978..2997)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3138..3160
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3154..3345)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     5700..5719
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5739..5816)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6022..6041
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg gggcagctgc tgcacggtgc gggcatctgt
      121 aagtggttca acgtgcgcat ggggttcggc ttcctgtcca tgaccgcccg cgccggggtc
      181 gcgctcgacc ccccagtgga tgtctttgtg caccagagta agctgcacat ggaagggttc
      241 cggagcttga aggagggtga ggcagtggag ttcaccttta agaagtcagc caagggtctg
      301 gaatccatcc gtgtcaccgg acctggtgga gtattctgta ttgggagtga gaggcggcca
      361 aaaggaaaga gcatgcagaa gcgcagatca aaaggagaca ggtgctacaa ctgtggaggt
      421 ctagatcatc atgccaagga atgcaagctg ccaccccagc ccaagaagtg ccacttctgc
      481 cagagcatca gccatatggt agcctcatgt ccgctgaagg cccagcaggg ccctagtgca
      541 cagggagggc atcatcacca tcaccactaa ctcgaggatc cggctgctaa caaagcccga
      601 aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc
      661 tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg atatcccgca
      721 agaggcccgg cagtaccggc ataaccaagc ctatgcctac agcatccagg gtgacggtgc
      781 cgaggatgac gatgagcgca ttgttagatt tcatacacgg tgcctgactg cgttagcaat
      841 ttaactgtga taaactaccg cattaaagct tatcgatgat aagctgtcaa acatgagaat
      901 tcttgaagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
      961 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     1021 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     1081 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     1141 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     1201 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     1261 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     1321 gttctgctat gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc
     1381 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     1441 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     1501 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     1561 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     1621 ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta
     1681 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     1741 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     1801 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     1861 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     1921 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     1981 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     2041 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     2101 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     2161 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     2221 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     2281 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     2341 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     2401 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     2461 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     2521 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     2581 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     2641 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     2701 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     2761 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     2821 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     2881 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     2941 ctgtgcggta tttcacaccg caatggtgca ctctcagtac aatctgctct gatgccgcat
     3001 agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca
     3061 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     3121 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     3181 acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc
     3241 tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct
     3301 gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta
     3361 agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat
     3421 acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc
     3481 ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa
     3541 tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat
     3601 aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac
     3661 cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc
     3721 gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct
     3781 caacgacagg agcacgatca tgcgcacccg tggccaggac ccaacgctgc ccgagatgcg
     3841 ccgcgtgcgg ctgctggaga tggcggacgc gatggatatg ttctgccaag ggttggtttg
     3901 cgcattcaca gttctccgca agaattgatt ggctccaatt cttggagtgg tgaatccgtt
     3961 agcgaggtgc cgccggcttc cattcaggtc gaggtggccc ggctccatgc accgcgacgc
     4021 aacgcgggga ggcagacaag gtatagggcg gcgcctacaa tccatgccaa cccgttccat
     4081 gtgctcgccg aggcggcata aatcgccgtg acgatcagcg gtccaatgat cgaagttagg
     4141 ctggtaagag ccgcgagcga tccttgaagc tgtccctgat ggtcgtcatc tacctgcctg
     4201 gacagcatgg cctgcaacgc gggcatcccg atgccgccgg aagcgagaag aatcataatg
     4261 gggaaggcca tccagcctcg cgtcgcgaac gccagcaaga cgtagcccag cgcgtcggcc
     4321 gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg
     4381 aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc
     4441 gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct
     4501 acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc
     4561 caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc
     4621 taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4681 aactgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     4741 ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc
     4801 accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga
     4861 aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg
     4921 tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt
     4981 gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc
     5041 agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct
     5101 atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc
     5161 gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga
     5221 tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc
     5281 tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg
     5341 gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga
     5401 ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg
     5461 ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc
     5521 agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt
     5581 gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt
     5641 ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg
     5701 gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc
     5761 tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg
     5821 atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag
     5881 gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc
     5941 cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg
     6001 gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt
     6061 ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg
     6121 aaat
//
