LOCUS       sequence-485487-        6547 bp    DNA     circular SYN 08-SEP-2025
DEFINITION  hygR selection nls-Scarlet marked rapid RMCE integration vector with
            an attP site for RMHE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6547)
  AUTHORS   Nonet ML
  TITLE     Creation and manipulation of bipartite expression transgenes in C. 
            elegans using phiC31 recombinase.
  JOURNAL   bioRxiv 2024.03.01.583017
REFERENCE   2  (bases 1 to 6547)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2024.03.01.583017"
COMMENT     Sequence Label: pHygRP2
FEATURES             Location/Qualifiers
     source          1..6547
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(100..117)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(271..859)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(351..370)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1030..1821,1822..1890))
                     /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     1653..1672
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1891..1995)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2021..2476)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2158..2179)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2370..2389
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    complement(2488..2521)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2638..2737
                     /label=phage phi-C31 attP
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attachment site of phage phi-C31"
     protein_bind    3226..3259
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             3272..3292
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    5091..5115
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(5116..6141)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(5116..6141)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     protein_bind    complement(6144..6168)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
ORIGIN
        1 atcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
       61 cggggagagg cggtttgcgt attgggcgaa ttccgcttcc tcgctcactg actcgctgcg
      121 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
      181 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
      241 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
      301 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
      361 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
      421 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
      481 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccat
      541 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
      601 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
      661 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
      721 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
      781 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
      841 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
      901 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
      961 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     1021 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     1081 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     1141 atctggcccc agtgctgcaa tgataccgcg agatccacgc tcaccggctc cagatttatc
     1201 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     1261 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     1321 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     1381 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     1441 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     1501 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     1561 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     1621 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     1681 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     1741 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     1801 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     1861 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     1921 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     1981 aataggggtt ccgcgcacat ttccccgaaa agtgccacct aaattgtaag cgttaatatt
     2041 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     2101 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     2161 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     2221 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     2281 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     2341 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     2401 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     2461 ccgctacagg gcgcgtcgtc ggagctcata acttcgtata gcatacatta tacgaagtta
     2521 tgcggccgct ctagaactag tggctgacat ccagtagacg gaggctaccg gtgtactgac
     2581 ggacacaccg aagccccggc ggcaaccctc agcggatgcc ccggggcttc acgttttccc
     2641 aggtcagaag cggttttcgg gagtagtgcc ccaactgggg taacctttga gttctctcag
     2701 ttgggggcgt agggtcgccg acatgacaca aggggttgtg accggggtgg acacgtacgc
     2761 gggtgcttac gaccgtcagt cgcgcgagcg cgacaattgt ctaaacattt tttgttgact
     2821 tacaataaat tgttgatata atttattgtg aaatgggtat tcttgaatga tttgatttta
     2881 acgaagagaa cgctgaaatc aactgaacac ctcgaattga gattttgggg attttatacc
     2941 aagttatcga cgacgcaaat ggtgcaaact ttatattttc aaatattctg gaagcattca
     3001 atggtctttt aagaaacgga caactccatt cttgatatgc aatgtttttt aaaagaaatc
     3061 tctttaatcg agacgggcgc atatagaacc taggtggcga agagcctgca ggtaaccacg
     3121 cgttgcatgc tcttccgtac ggcgacgtaa cgtgagatag gcggaacccg ggaagttcct
     3181 atactatttg aagaatagga acttctgcag gaattcgata tcggtgaagt tcctattctc
     3241 tagaaagtat aggaacttca aaatgggatc cccaaagaag aagcgtaagg tgtcgatggt
     3301 ttccaaggga gaggccgtca tcaaggagtt catgcgtttc aaggtccaca tggagggatc
     3361 catgaacgga cacgagttcg agatcgaggg agagggagag ggacgtccat acgagggaac
     3421 ccaaaccgcc aagctcaagg tcaccaaggt aagtttaaac atatatatac taactaaccc
     3481 tgattattta aattttcagg gaggaccact cccattctcc tgggacatcc tctccccaca
     3541 attcatgtac ggatcccgtg ccttcaccaa gcacccagcc gacatcccag actactacaa
     3601 gcaatccttc ccagagggat tcaagtggga gcgtgtcatg aacttcgagg acggaggagc
     3661 cgtcaccgtc acccaagaca cctccctcga ggacggaacc ctcatctaca aggtaagttt
     3721 aaacagttcg gtactaacta accatacata tttaaatttt caggtcaagc tccgtggaac
     3781 caacttccca ccagacggac cagtcatgca aaagaagacc atgggatggg aggcctccac
     3841 cgagcgtctc tacccagagg acggagtcct caagggagac atcaagatgg ccctccgtct
     3901 caaggacgga ggacgttacc tcgccgactt caaggtaagt ttaaacatga ttttactaac
     3961 taactaatct gatttaaatt ttcagaccac ctacaaggcc aagaagccag tccaaatgcc
     4021 aggagcctac aacgtcgacc gtaagctcga catcacctcc cacaacgagg actacaccgt
     4081 cgtcgagcaa tacgagcgtt ccgagggacg tcactccacc ggaggaatgg acgagctcta
     4141 caaggcctcc cgtcgtcgta aggccaaccc aaccaagctc tccgagaacg ccaagaagct
     4201 cgccaagtag atcaactgct ctaattccat ctcgtttctc tcttttcttt caatctcttt
     4261 acccgtttct cttcccctac gtcccccaaa cctaccactg cacagtaaca catacacatt
     4321 caacgtgtca caataaaagc ccatcaacac cttctctgtt gttttttttc ttcaaatgca
     4381 tttttttcac tttgtttaat aatttttacg catcggattt gtaggatagt ggaaatttgt
     4441 ccttcaatgc ttcaattgat tcgacacttc tcaatatttt tgatcaataa accattgatt
     4501 ttctatttag aagaagatca ctgtaaataa caccaaaaac atcacagcaa aaaaactccc
     4561 actcaattat attactgttt ttattgaaac tagaataaat ttgaaaatct tcaaagcata
     4621 cacaattaaa aaaaacgaaa aaagaaaaag tcccgttgat tctcaaactt aaatttttaa
     4681 acttgtctga accatgttgt aagtttgtaa aagtttccca gttgtcctcg aacaggtcta
     4741 aaaattgcgg tcataaactg aaacgtaaca tatgataagg tattttgtgt gcggtttttt
     4801 ctatgatgtt tcaaaaaaat ttatcagaag taaaaaacat aagaagtgta cacagaacat
     4861 tgtgtagatt ttgggataga attattatta atggggggag ggagcacaat ttttttcgac
     4921 tttttattac gaattaattc tatttttgaa tctacacaat ttcattgtta gaggtgactt
     4981 aaaagaagct aaaaaacaaa gaaattaaga gaagtttttt gataataaca aaaatagggg
     5041 gtgggagcac agggagaaag agcatgtagg gatgttgaag agtaattgga cccactttgt
     5101 acaagaaagc tgggtctatt cctttgccct cggacgagtg ctggggcgtc ggtttccact
     5161 atcggcgagt acttctacac agccatcggt ccagacggcc gcgcttctgc gggcgatttg
     5221 tgtacgcccg acagtcccgg ctccggatcg gacgattgcg tcgcatcgac cctgcgccca
     5281 agctgcatca tcgaaattgc cgtcaaccaa gctctgatag agttggtcaa gaccaatgcg
     5341 gagcatatac gcccggagcc gcggcgatcc tgcaagctcc ggatgcctcc gctcgaagta
     5401 gcgcgtctgc tgctccatac aagccaacca cggcctccag aagaagatgt tggcgacctc
     5461 gtattgggaa tccccgaaca tcgcctcgct ccagtcaatg accgctgtta tgcggccatt
     5521 gtccgtcagg acattgttgg agccgaaatc cgcgtgcacg aggtgccgga cttcggggca
     5581 gtcctcggcc caaagcatca gctcatcgag agcctgcgcg acggacgcac tgacggtgtc
     5641 gtccatcaca gtttgccagt gatacacatg gggatcagca atcgcgcata tgaaatcacg
     5701 ccatgtagtg tattgaccga ttccttgcgg tccgaatggg ccgaacccgc tcgtctggct
     5761 aagatcggcc gcagcgatcg catccatggc ctccgcgacc ggctgcagaa cagcgggcag
     5821 ttcggtttca ggcaggtctt gcaacgtgac accctgtgca cggcgggaga tgcaataggt
     5881 caggctctcg ctgaattccc caatgtcaag cacttccgga atcgggagcg cggccgatgc
     5941 aaagtgccga taaacataac gatctttgta gaaaccatcg gcgcagctat ttacccgcag
     6001 gacatatcca cgccctccta catcgaagct gaaagcacga gattcttcgc cctccgagag
     6061 ctgcatcagg tcggagacgc tgtcgaactt ttcgatcaga aacttctcga cagacgtcgc
     6121 ggtgagttca ggctttttca tggagcctgc ttttttgtac aaacttgcgg caccgcctga
     6181 catattacct taaaattcaa aaattaattt cagatcagtc ataaccaaga aaacaaaata
     6241 aacgttagga cagaaaattc aaattaatta aaaattatgc tgaataagaa aattacatgc
     6301 aaagccttat tttcctcata gaaacagttg aataacatat tttttaattg aaaacatagc
     6361 aaaattcgtg gtaaaatttg tcaaaacctt ggaaaaaatc gaaaaaagaa catttcccga
     6421 tgcattttcc tatttctcgt tgccgacaac gtataaacgt tgaatactca gtcaacgaat
     6481 ttggctgctg gcttggactc tcgcacggaa gagacgcgtg tgtagattta cgacgaaagc
     6541 aaaaatg
//
