LOCUS       sequence-441836-        6523 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses R141S K142S LoaP double mutant.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6523)
  AUTHORS   Elghondakly A, Jermain MD, Winkler WC, Ferre-D'Amare AR
  TITLE     Major-groove sequence-specific RNA recognition by LoaP, a paralog of
            transcription elongation factor NusG.
  JOURNAL   Structure. 2024 Jun 28:S0969-2126(24)00219-3. doi: 
            10.1016/j.str.2024.06.001.
  PUBMED    38959899
REFERENCE   2  (bases 1 to 6523)
  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"; journalName: "Structure. 
            2024 Jun 28:S0969-2126(24)00219-3. doi: 10.1016/j.str.2024.06.001."
COMMENT     Sequence Label: pQA-LP580
FEATURES             Location/Qualifiers
     source          1..6523
                     /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"
     misc_feature    775..914
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(840..859)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1000..1022
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1016..1207)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     3562..3581
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3601..3678)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3884..3903
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3991..4010
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3991..4009
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4010..4034
                     /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             4049..4071
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4084..4113
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             4129..4143
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     primer_bind     complement(5007..5025)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5021..5068
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(5243..5248,5249..5265,5266..5271))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(5302..5320)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5388..5492
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5493..5561,5562..6353)
                     /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(5711..5730)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, 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 gaagcggaag agcgcctgat
      781 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcaatggtg cactctcagt
      841 acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact
      901 gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
      961 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     1021 ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt
     1081 cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca
     1141 gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt
     1201 tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga
     1261 aacgagagag gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac
     1321 gttgtgaggg taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg
     1381 gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc
     1441 ctgcgatgca gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt
     1501 acgaaacacg gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc
     1561 agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac
     1621 cccgccagcc tagccgggtc ctcaacgaca ggagcacgat catgcgcacc cgtggccagg
     1681 acccaacgct gcccgagatg cgccgcgtgc ggctgctgga gatggcggac gcgatggata
     1741 tgttctgcca agggttggtt tgcgcattca cagttctccg caagaattga ttggctccaa
     1801 ttcttggagt ggtgaatccg ttagcgaggt gccgccggct tccattcagg tcgaggtggc
     1861 ccggctccat gcaccgcgac gcaacgcggg gaggcagaca aggtataggg cggcgcctac
     1921 aatccatgcc aacccgttcc atgtgctcgc cgaggcggca taaatcgccg tgacgatcag
     1981 cggtccaatg atcgaagtta ggctggtaag agccgcgagc gatccttgaa gctgtccctg
     2041 atggtcgtca tctacctgcc tggacagcat ggcctgcaac gcgggcatcc cgatgccgcc
     2101 ggaagcgaga agaatcataa tggggaaggc catccagcct cgcgtcgcga acgccagcaa
     2161 gacgtagccc agcgcgtcgg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg
     2221 tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc
     2281 aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca
     2341 gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc
     2401 gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg
     2461 catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc
     2521 actgcccgct ttccagtcgg gaaactgtcg tgccagctgc attaatgaat cggccaacgc
     2581 gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca ccagtgagac
     2641 gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac
     2701 gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg ggatataaca
     2761 tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa cgcgcagccc
     2821 ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc
     2881 agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg acatggcact
     2941 ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat atttatgcca
     3001 gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca gcgcgatttg
     3061 ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt catgggagaa
     3121 aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg gaacattagt
     3181 gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa tgatcagccc
     3241 actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga cgccgcttcg
     3301 ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc
     3361 gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga
     3421 ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc
     3481 cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga
     3541 aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac
     3601 attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt
     3661 gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc tgcattagga
     3721 agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca
     3781 aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac
     3841 aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat
     3901 aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga
     3961 ggatcgagat cgatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag
     4021 cggataacaa ttcccctcta gaaataattt tgtttaactt taagaaggag atataccatg
     4081 ggccatcatc atcatcatca tcatcatcat cacagcagcg gccatatcga cgacgacgac
     4141 aagcatagcc acattaactt aaaagtaaag gggcaagatg ggaacgaggt gttcttccgt
     4201 atcaagcgca gtacacagtt aaaaaagctg atgaatgctt attgtgatcg tcaatctgtc
     4261 gacatgacag caattgcatt ccttttcgat gggcgtcgtt tgcgtgcaga acaaacaccc
     4321 gacgaattag agatggaaga cggcgatgaa attgacgcga tgcttcatca gacaggaggt
     4381 gccggaacaa tgaagaaatg gtacgtgatc ttcacgcgct caggttatga aaacaaagtt
     4441 cgtgacatca tcgaaaattg cttcaaagag gaagttaaat tactgattcc taagcgcaaa
     4501 attattgagc gtgtcaaggg ccaacccgtt gaaaaaatca aattgctgtt tcctggttat
     4561 gtgtttgtaa atgcagaaat gtctgatgat ctttattaca agattagtga agtactgaag
     4621 cgcggtatct tcttgaagga gggaaagcgc cccgctttcg ttaaagaaga agaaatgaaa
     4681 attattttaa gtttgacaaa gaattcggat cttattgatc ttagtaaggg tattatggag
     4741 ggagaacgcg taaaaatcat cgaagggccg ttaaaagggt acgaaggcct tattaaaaag
     4801 attgataaaa gcagcaagcg tgctaaggtt atctttagca ttgcagggga attaaaatct
     4861 gtggatttgg cgatcgaggt tatggagaac gtcagcgagc agcagcgttc cttggtctac
     4921 gcgtgttgac atatgctcga ggatcctgat taattgagat ccggctgcta acaaagcccg
     4981 aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc
     5041 ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg gatatcccgc
     5101 aagaggcccg gcagtaccgg cataaccaag cctatgccta cagcatccag ggtgacggtg
     5161 ccgaggatga cgatgagcgc attgttagat ttcatacacg gtgcctgact gcgttagcaa
     5221 tttaactgtg ataaactacc gcattaaagc ttatcgatga taagctgtca aacatgagaa
     5281 ttaattcttg aagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     5341 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     5401 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     5461 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     5521 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     5581 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     5641 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     5701 ttaaagttct gctatgtggc gcggtattat cccgtgttga cgccgggcaa gagcaactcg
     5761 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     5821 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     5881 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     5941 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     6001 ccataccaaa cgacgagcgt gacaccacga tgcctgcagc aatggcaaca acgttgcgca
     6061 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     6121 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     6181 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     6241 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     6301 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     6361 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     6421 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     6481 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttc
//
