LOCUS       sequence-485986-        6577 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  hygR selection nls-Scarlet marked rapid RMCE integration vector with
            an attB site for RMHE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6577)
  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 6577)
  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: "bioRxiv 
            2024.03.01.583017"
COMMENT     Sequence Label: pHygRB2
FEATURES             Location/Qualifiers
     source          1..6577
                     /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    2901..2970
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     protein_bind    complement(3202..3235)
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     protein_bind    3256..3289
                     /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             3302..3322
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    5121..5145
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(5146..6171)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(5146..6171)
                     /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(6174..6198)
                     /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 gaggctcaat tgaaagtaaa
     2581 tattccctgt gtttaattgt tattgttgtg gtggataata gatatttaat ctgccaaaat
     2641 gtgttctttt tacatagctg atcaatttcc agatatatcg agacttgatg tcattcctgg
     2701 atgaatcgtc taatggtgtt aaacctgatt tgattagttg tctctttgtc gtattagagt
     2761 ggatgagttc atccagttgt tgaaaaaatg ttcttattgt gaagaaaaac taattcgtag
     2821 aacctaggtg gcgaagagcc tgcaggtaac cacgcgttgc atgctcttcc gtacgtaccg
     2881 tcgacgatgt aggtcacggt ctcgaagccg cggtgcgggt gccagggcgt gcccttgggc
     2941 tccccgggcg cgtactccac ctcacccatc tggtccatca tgatgaacgg gtcgaggtgg
     3001 cggtagttga tcccggcgaa cgcgcggcgc accgggaagc cctcgccctc gaaaccgctg
     3061 ggcgcggtgg tcacggtgag cacgggacgt gcgacggcgt cggcgggtgc ggatacgcgg
     3121 ggcagcgtca gcgggttctc gacggtcacg gcgggcatgt cgacacttaa gcgacgtaac
     3181 gtgagatagg cggaacaccg ggaagttcct atactatttg aagaatagga acttctgcag
     3241 gaattcgata tcggtgaagt tcctattctc tagaaagtat aggaacttca aaatgggatc
     3301 cccaaagaag aagcgtaagg tgtcgatggt ttccaaggga gaggccgtca tcaaggagtt
     3361 catgcgtttc aaggtccaca tggagggatc catgaacgga cacgagttcg agatcgaggg
     3421 agagggagag ggacgtccat acgagggaac ccaaaccgcc aagctcaagg tcaccaaggt
     3481 aagtttaaac atatatatac taactaaccc tgattattta aattttcagg gaggaccact
     3541 cccattctcc tgggacatcc tctccccaca attcatgtac ggatcccgtg ccttcaccaa
     3601 gcacccagcc gacatcccag actactacaa gcaatccttc ccagagggat tcaagtggga
     3661 gcgtgtcatg aacttcgagg acggaggagc cgtcaccgtc acccaagaca cctccctcga
     3721 ggacggaacc ctcatctaca aggtaagttt aaacagttcg gtactaacta accatacata
     3781 tttaaatttt caggtcaagc tccgtggaac caacttccca ccagacggac cagtcatgca
     3841 aaagaagacc atgggatggg aggcctccac cgagcgtctc tacccagagg acggagtcct
     3901 caagggagac atcaagatgg ccctccgtct caaggacgga ggacgttacc tcgccgactt
     3961 caaggtaagt ttaaacatga ttttactaac taactaatct gatttaaatt ttcagaccac
     4021 ctacaaggcc aagaagccag tccaaatgcc aggagcctac aacgtcgacc gtaagctcga
     4081 catcacctcc cacaacgagg actacaccgt cgtcgagcaa tacgagcgtt ccgagggacg
     4141 tcactccacc ggaggaatgg acgagctcta caaggcctcc cgtcgtcgta aggccaaccc
     4201 aaccaagctc tccgagaacg ccaagaagct cgccaagtag atcaactgct ctaattccat
     4261 ctcgtttctc tcttttcttt caatctcttt acccgtttct cttcccctac gtcccccaaa
     4321 cctaccactg cacagtaaca catacacatt caacgtgtca caataaaagc ccatcaacac
     4381 cttctctgtt gttttttttc ttcaaatgca tttttttcac tttgtttaat aatttttacg
     4441 catcggattt gtaggatagt ggaaatttgt ccttcaatgc ttcaattgat tcgacacttc
     4501 tcaatatttt tgatcaataa accattgatt ttctatttag aagaagatca ctgtaaataa
     4561 caccaaaaac atcacagcaa aaaaactccc actcaattat attactgttt ttattgaaac
     4621 tagaataaat ttgaaaatct tcaaagcata cacaattaaa aaaaacgaaa aaagaaaaag
     4681 tcccgttgat tctcaaactt aaatttttaa acttgtctga accatgttgt aagtttgtaa
     4741 aagtttccca gttgtcctcg aacaggtcta aaaattgcgg tcataaactg aaacgtaaca
     4801 tatgataagg tattttgtgt gcggtttttt ctatgatgtt tcaaaaaaat ttatcagaag
     4861 taaaaaacat aagaagtgta cacagaacat tgtgtagatt ttgggataga attattatta
     4921 atggggggag ggagcacaat ttttttcgac tttttattac gaattaattc tatttttgaa
     4981 tctacacaat ttcattgtta gaggtgactt aaaagaagct aaaaaacaaa gaaattaaga
     5041 gaagtttttt gataataaca aaaatagggg gtgggagcac agggagaaag agcatgtagg
     5101 gatgttgaag agtaattgga cccactttgt acaagaaagc tgggtctatt cctttgccct
     5161 cggacgagtg ctggggcgtc ggtttccact atcggcgagt acttctacac agccatcggt
     5221 ccagacggcc gcgcttctgc gggcgatttg tgtacgcccg acagtcccgg ctccggatcg
     5281 gacgattgcg tcgcatcgac cctgcgccca agctgcatca tcgaaattgc cgtcaaccaa
     5341 gctctgatag agttggtcaa gaccaatgcg gagcatatac gcccggagcc gcggcgatcc
     5401 tgcaagctcc ggatgcctcc gctcgaagta gcgcgtctgc tgctccatac aagccaacca
     5461 cggcctccag aagaagatgt tggcgacctc gtattgggaa tccccgaaca tcgcctcgct
     5521 ccagtcaatg accgctgtta tgcggccatt gtccgtcagg acattgttgg agccgaaatc
     5581 cgcgtgcacg aggtgccgga cttcggggca gtcctcggcc caaagcatca gctcatcgag
     5641 agcctgcgcg acggacgcac tgacggtgtc gtccatcaca gtttgccagt gatacacatg
     5701 gggatcagca atcgcgcata tgaaatcacg ccatgtagtg tattgaccga ttccttgcgg
     5761 tccgaatggg ccgaacccgc tcgtctggct aagatcggcc gcagcgatcg catccatggc
     5821 ctccgcgacc ggctgcagaa cagcgggcag ttcggtttca ggcaggtctt gcaacgtgac
     5881 accctgtgca cggcgggaga tgcaataggt caggctctcg ctgaattccc caatgtcaag
     5941 cacttccgga atcgggagcg cggccgatgc aaagtgccga taaacataac gatctttgta
     6001 gaaaccatcg gcgcagctat ttacccgcag gacatatcca cgccctccta catcgaagct
     6061 gaaagcacga gattcttcgc cctccgagag ctgcatcagg tcggagacgc tgtcgaactt
     6121 ttcgatcaga aacttctcga cagacgtcgc ggtgagttca ggctttttca tggagcctgc
     6181 ttttttgtac aaacttgcgg caccgcctga catattacct taaaattcaa aaattaattt
     6241 cagatcagtc ataaccaaga aaacaaaata aacgttagga cagaaaattc aaattaatta
     6301 aaaattatgc tgaataagaa aattacatgc aaagccttat tttcctcata gaaacagttg
     6361 aataacatat tttttaattg aaaacatagc aaaattcgtg gtaaaatttg tcaaaacctt
     6421 ggaaaaaatc gaaaaaagaa catttcccga tgcattttcc tatttctcgt tgccgacaac
     6481 gtataaacgt tgaatactca gtcaacgaat ttggctgctg gcttggactc tcgcacggaa
     6541 gagacgcgtg tgtagattta cgacgaaagc aaaaatg
//
