LOCUS       sequence-372779-        4779 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses E. Coli RNase III under control of pBAD promoter with 
            native 5' untranslated region and constitutively expresses 
            gUTR-129-GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4779)
  AUTHORS   Weeks R, Ostermeier M
  TITLE     Fitness and functional landscapes of the E. coli RNase III gene rnc.
  JOURNAL   Mol Biol Evol. 2023 Feb 27:msad047. doi: 10.1093/molbev/msad047.
  PUBMED    36848192
REFERENCE   2  (bases 1 to 4779)
  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: "Mol Biol 
            Evol. 2023 Feb 27:msad047. doi: 10.1093/molbev/msad047."
COMMENT     Sequence Label: pRnc2-gGFP
FEATURES             Location/Qualifiers
     source          1..4779
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        16..50
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     misc_RNA        200..275
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             296..325
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             341..358
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(405..1283)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        1310..1594
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     1517..1536
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     primer_bind     complement(2480..2497)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2650..2696
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(2749..2766)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2920..3508)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3000..3019)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3679..4470,4471..4539))
                     /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     4302..4321
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4540..4644)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4712..4730
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(4734..4777)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
ORIGIN
        1 gaattctaaa gatctttgac agctagctca gtcctaggta taatactagt gaagtttaag
       61 gttggcacct ccaggttgcc acggcacacg aaacagcgtt ggtccccata taccggtaaa
      121 ctgaaactgc agcgaagcag ttagcagaac catgtatatc aggtctgttt cgtgtgctgc
      181 atctaattca acaagaattg ttttagagct agaaatagca agttaaaata aggctagtcc
      241 gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt ttgaagcttg ggcccgaaca
      301 aaaactcatc tcagaagagg atctgaatag cgccgtcgac catcatcatc atcatcattg
      361 agtttaaacg gtctccagcc gtcaattgtc tgattcgtta ccaattatga caacttgacg
      421 gctacatcat tcactttttc ttcacaaccg gcacggaact cgctcgggct ggccccggtg
      481 cattttttaa atacccgcga gaaatagagt tgatcgtcaa aaccaacatt gcgaccgacg
      541 gtggcgatag gcatccgggt ggtgctcaaa agcagcttcg cctggctgat acgttggtcc
      601 tcgcgccagc ttaagacgct aatccctaac tgctggcgga aaagatgtga cagacgcgac
      661 ggcgacaagc aaacatgctg tgcgacgctg gcgatatcaa aattgctgtc tgccaggtga
      721 tcgctgatgt actgacaagc ctcgcgtacc cgattatcca tcggtggatg gagcgactcg
      781 ttaatcgctt ccatgcgccg cagtaacaat tgctcaagca gatttatcgc cagcagctcc
      841 gaatagcgcc cttccccttg cccggcgtta atgatttgcc caaacaggtc gctgaaatgc
      901 ggctggtgcg cttcatccgg gcgaaagaac cccgtattgg caaatattga cggccagtta
      961 agccattcat gccagtaggc gcgcggacga aagtaaaccc actggtgata ccattcgcga
     1021 gcctccggat gacgaccgta gtgatgaatc tctcctggcg ggaacagcaa aatatcaccc
     1081 ggtcggcaaa caaattctcg tccctgattt ttcaccaccc cctgaccgcg aatggtgaga
     1141 ttgagaatat aacctttcat tcccagcggt cggtcgataa aaaaatcgag ataaccgttg
     1201 gcctcaatcg gcgttaaacc cgccaccaga tgggcattaa acgagtatcc cggcagcagg
     1261 ggatcatttt gcgcttcagc catacttttc atactcccgc cattcagaga agaaaccaat
     1321 tgtccatatt gcatcagaca ttgccgtcac tgcgtctttt actggctctt ctcgctaacc
     1381 aaaccggtaa ccccgcttat taaaagcatt ctgtaacaaa gcgggaccaa agccatgaca
     1441 aaaacgcgta acaaaagtgt ctataatcac ggcagaaaag tccacattga ttatttgcac
     1501 ggcgtcacac tttgctatgc catagcattt ttatccataa gattagcgga tcctacctga
     1561 cgctttttat cgcaactctc tactgtttct ccatgaagtt taaggttggc acctccaggt
     1621 tgccacggca cacgaaacag cgttggtccc catataccgg taaactgaaa ctgcagcgaa
     1681 gcagttagca gaaccatgta tatcaggtct gtttcgtgtg ctgaattgtt gacgcattta
     1741 tttattggta tcgcatgaac cccatcgtaa ttaatcggct tcaacggaag ctgggctaca
     1801 cttttaatca tcaggaactg ttgcagcagg cattaactca tcgtagtgcc agcagtaaac
     1861 ataacgagcg tttagaattt ttaggcgact ctattctgag ctacgttatc gccaatgcgc
     1921 tttatcaccg tttccctcgt gtggatgaag gcgatatgag ccggatgcgc gccacgctgg
     1981 tccgtggcaa tacgctggcg gaactggcgc gcgaatttga gttaggcgag tgcttacgtt
     2041 tagggccagg tgaacttaaa agcggtggat ttcgtcgtga gtcaattctc gccgacaccg
     2101 tcgaagcatt aattggtggc gtattcctcg acagtgatat tcaaaccgtc gagaaattaa
     2161 tcctcaactg gtatcaaact cgtttggacg aaattagccc aggcgataaa caaaaagatc
     2221 cgaaaacgcg cttgcaagaa tatttgcagg gtcgccatct gccgctgccg acttatctgg
     2281 tagtccaggt acgtggcgaa gcgcacgatc aggaatttac tatccactgc caggtcagcg
     2341 gcctgagtga accggtggtt ggcacaggtt caagccgtcg taaggctgag caggctgccg
     2401 ccgaacaggc gttgaaaaaa ctggagctgg aatgagagct caagcttggc tgttttggcg
     2461 gatgagagaa gattttcagc ctgatacaga ttaaatcaga acgcagaagc ggtctgataa
     2521 aacagaattt gcctggcggc agtagcgcgg tggtcccacc tgaccccatg ccgaactcag
     2581 aagtgaaacg ccgtagcgcc gatggtagtg tggggtctcc ccatgcgaga gtagggaact
     2641 gccaggcatc aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt
     2701 tgtttgtcgg tgaactggat ccttactcga gtctagactg caggcttcct cgctcactga
     2761 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     2821 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     2881 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccacaggc tccgcccccc
     2941 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     3001 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     3061 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     3121 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     3181 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     3241 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     3301 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     3361 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     3421 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     3481 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     3541 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     3601 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     3661 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     3721 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     3781 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     3841 agatttatca gcaataaacc agccagctgg aagggccgag cgcagaagtg gtcctgcaac
     3901 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     3961 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     4021 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     4081 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     4141 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     4201 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     4261 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     4321 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     4381 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     4441 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     4501 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     4561 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     4621 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga
     4681 aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc agaatttcag
     4741 ataaaaaaaa tccttagctt tcgctaagga tgatttctg
//
