LOCUS       sequence-174747-        5472 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pET LIC cloning vector for yORF-mRuby2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5472)
  TITLE     C-terminal fluorescent fusion vectors for E. coli expression
REFERENCE   2  (bases 1 to 5472)
  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"
COMMENT     Sequence Label: pET mRuby2 LIC cloning vector (u-mRuby2)
FEATURES             Location/Qualifiers
     source          1..5472
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      195..783
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     684..703
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     937..954
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    969..1108
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(1034..1053)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1194..1216
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1210..1401)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     2859..2878
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2962..2981
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2962..2980
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             3069..3091
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3132..3839
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="VSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTMR
                     IKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGGV
                     VTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTHM
                     ALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAVA
                     KFAGLGGGMDELYK"
     primer_bind     complement(3963..3981)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3977..4024
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(4390..4395,4396..4412,4413..4418))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(4445..4463)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4531..4635
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4636..4704,4705..5472,1..24)
                     /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(4854..4873)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
       61 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
      121 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
      181 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
      241 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
      301 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
      361 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
      421 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
      481 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
      541 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
      601 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
      661 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
      721 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
      781 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
      841 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
      901 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
      961 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat
     1021 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct
     1081 atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc
     1141 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     1201 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag
     1261 ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc
     1321 gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc
     1381 ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt
     1441 aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc
     1501 ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag
     1561 aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg
     1621 tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg
     1681 cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc
     1741 agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta
     1801 accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg
     1861 cacccgtggc caggacccaa cgctgcccga gatgcgccgc gtgcggctgc tggagatggc
     1921 ggacgcgatg gatatgttct gccaagggtt ggtttgcgca ttcacagttc tccgcaagaa
     1981 ttgattggct ccaattcttg gagtggtgaa tccgttagcg aggtgccgcc ggcttccatt
     2041 caggtcgagg tggcccggct ccatgcaccg cgacgcaacg cggggaggca gacaaggtat
     2101 agggcggcgc ctacaatcca tgccaacccg ttccatgtgc tcgccgaggc ggcataaatc
     2161 gccgtgacga tcagcggtcc aatgatcgaa gttaggctgg taagagccgc gagcgatcct
     2221 tgaagctgtc cctgatggtc gtcatctacc tgcctggaca gcatggcctg caacgcgggc
     2281 atcccgatgc cgccggaagc gagaagaatc ataatgggga aggccatcca gcctcgcgtc
     2341 gcgaacgcca gcaagacgta gcccagcgcg tcggccgcca tgccggcgat aatggcctgc
     2401 ttctcgccga aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag
     2461 attccgaata ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg
     2521 ccgaaaatga cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca
     2581 gtcataagtg cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg
     2641 aaggctctca agggcatcgg tcgacgctct cccttatgcg actcctgcat taggaagcag
     2701 cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag
     2761 atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg
     2821 ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg
     2881 ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc
     2941 gagatctcga tcccgcgaaa ttaatacgac tcactatagg gagaccacaa cggtttccct
     3001 ctagtgccgg ctccggagag ctctttaatt aagcggccgc cctgcaggac tcgagttcta
     3061 gaaataattt tgtttaactt taagaaggag atatagatat cctctcatcc tccaacggca
     3121 gcagcggatc cgtgtctaag ggcgaagagc tgatcaagga aaatatgcgt atgaaggtgg
     3181 tcatggaagg ttcggtcaac ggccaccaat tcaaatgcac aggtgaagga gaaggcaatc
     3241 cgtacatggg aactcaaacc atgaggatca aagtcatcga gggaggaccc ctgccatttg
     3301 cctttgacat tcttgccacg tcgttcatgt atggcagccg tacttttatc aagtacccga
     3361 aaggcattcc tgatttcttt aaacagtcct ttcctgaggg ttttacttgg gaaagagtta
     3421 cgagatacga agatggtgga gtcgtcaccg tcatgcagga caccagcctt gaggatggct
     3481 gtctcgttta ccacgtccaa gtcagagggg taaactttcc ctccaatggt cccgtgatgc
     3541 agaagaagac caagggttgg gagcctaata cagagatgat gtatccagca gatggtggtc
     3601 tgaggggata cactcatatg gcactgaaag ttgatggtgg tggccatctg tcttgctctt
     3661 tcgtaacaac ttacaggtca aaaaagaccg tcgggaacat caagatgccc ggtatccatg
     3721 ccgttgatca ccgcctggaa aggttagagg aaagtgacaa tgaaatgttc gtagtacaac
     3781 gcgaacacgc agttgccaag ttcgccgggc ttggtggtgg gatggacgag ctgtacaagt
     3841 agtaaccaac tccataagga tccgcgatcg cggcgcgcca cctggtggcc ggccggtacc
     3901 acgcgtgcgc gctgatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc
     3961 caccgctgag caataactag cataacccct tggggcctct aaacgggtct tgaggggttt
     4021 tttgctgaaa ggaggaacta tatccggaca tccacaggac gggtgtggtc gccatgatcg
     4081 cgtagtcgat agtggctcca agtagcgaag cgagcaggac tgggcggcgg ccaaagcggt
     4141 cggacagtgc tccgagaacg ggtgcgcata gaaattgcat caacgcatat agcgctagca
     4201 gcacgccata gtgactggcg atgctgtcgg aatggacgac atcccgcaag aggcccggca
     4261 gtaccggcat aaccaagcct atgcctacag catccagggt gacggtgccg aggatgacga
     4321 tgagcgcatt gttagatttc atacacggtg cctgactgcg ttagcaattt aactgtgata
     4381 aactaccgca ttaaagctta tcgatgataa gctgtcaaac atgagaattc ttgaagacga
     4441 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     4501 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     4561 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataacat
     4621 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     4681 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     4741 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     4801 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     4861 ggcgcggtat tatcccgtgt tgacgccggg caagagcaac tcggtcgccg catacactat
     4921 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     4981 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     5041 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     5101 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     5161 cgtgacacca cgatgcctgc agcaatggca acaacgttgc gcaaactatt aactggcgaa
     5221 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     5281 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     5341 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     5401 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     5461 gctgagatag gt
//
