LOCUS       sequence-485981-        6592 bp    DNA     circular SYN 16-SEP-2025
DEFINITION  hygR selection nls-GFP 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 6592)
  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 6592)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 16, 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: pHyGB6
FEATURES             Location/Qualifiers
     source          1..6592
                     /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    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    5136..5160
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(5161..6186)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(5161..6186)
                     /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(6189..6213)
                     /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 tgtcgatgag taaaggagaa gaattgttca ctggagttgt
     3361 cccaatcctc gtcgagctcg acggagacgt caacggacac aagttctccg tctccggaga
     3421 gggagaggga gacgccacct acggaaagct caccctcaag ttcatctgca ccaccggaaa
     3481 gctcccagtc ccatggccaa ccctcgtcac caccttctgc tacggagtcc aatgcttctc
     3541 ccgttaccca gaccacatga agcgtcacga cttcttcaag tccgccatgc cagagggata
     3601 cgtccaagag cgtaccatct tcttcaaggt aagtttaaac atatatatac taactactga
     3661 ttatttaaat tttcaggacg acggaaacta caagacccgt gccgaggtca agttcgaggg
     3721 agacaccctc gtcaaccgta tcgagctcaa ggtaagttta aacagttcgg tactaactaa
     3781 ccatacatat ttaaattttc agggaatcga cttcaaggag gacggaaaca tcctcggaca
     3841 caagctcgag tacaactaca actcccacaa cgtctacatc atggccgaca agcaaaagaa
     3901 cggaatcaag gtcaacttca aggtaagttt aaacatgatt ttactaacta actaatctga
     3961 tttaaatttt cagatccgtc acaacatcga ggacggatcc gtccaactcg ccgaccacta
     4021 ccaacaaaac accccaatcg gagacggacc agtcctcctc ccagacaacc actacctctc
     4081 cacccaatcc gccctctcca aggacccaaa cgagaagcgt gaccacatgg tcctcctcga
     4141 gttcgtcacc gccgccggaa tcacccacgg aatggacgag ctctacaagg cctcccgtcg
     4201 tcgtaaggcc aacccaacca agctctccga gaacgccaag aagctcgcca agtagatcaa
     4261 ctgctctaat tccatctcgt ttctctcttt tctttcaatc tctttacccg tttctcttcc
     4321 cctacgtccc ccaaacctac cactgcacag taacacatac acattcaacg tgtcacaata
     4381 aaagcccatc aacaccttct ctgttgtttt ttttcttcaa atgcattttt ttcactttgt
     4441 ttaataattt ttacgcatcg gatttgtagg atagtggaaa tttgtccttc aatgcttcaa
     4501 ttgattcgac acttctcaat atttttgatc aataaaccat tgattttcta tttagaagaa
     4561 gatcactgta aataacacca aaaacatcac agcaaaaaaa ctcccactca attatattac
     4621 tgtttttatt gaaactagaa taaatttgaa aatcttcaaa gcatacacaa ttaaaaaaaa
     4681 cgaaaaaaga aaaagtcccg ttgattctca aacttaaatt tttaaacttg tctgaaccat
     4741 gttgtaagtt tgtaaaagtt tcccagttgt cctcgaacag gtctaaaaat tgcggtcata
     4801 aactgaaacg taacatatga taaggtattt tgtgtgcggt tttttctatg atgtttcaaa
     4861 aaaatttatc agaagtaaaa aacataagaa gtgtacacag aacattgtgt agattttggg
     4921 atagaattat tattaatggg gggagggagc acaatttttt tcgacttttt attacgaatt
     4981 aattctattt ttgaatctac acaatttcat tgttagaggt gacttaaaag aagctaaaaa
     5041 acaaagaaat taagagaagt tttttgataa taacaaaaat agggggtggg agcacaggga
     5101 gaaagagcat gtagggatgt tgaagagtaa ttggacccac tttgtacaag aaagctgggt
     5161 ctattccttt gccctcggac gagtgctggg gcgtcggttt ccactatcgg cgagtacttc
     5221 tacacagcca tcggtccaga cggccgcgct tctgcgggcg atttgtgtac gcccgacagt
     5281 cccggctccg gatcggacga ttgcgtcgca tcgaccctgc gcccaagctg catcatcgaa
     5341 attgccgtca accaagctct gatagagttg gtcaagacca atgcggagca tatacgcccg
     5401 gagccgcggc gatcctgcaa gctccggatg cctccgctcg aagtagcgcg tctgctgctc
     5461 catacaagcc aaccacggcc tccagaagaa gatgttggcg acctcgtatt gggaatcccc
     5521 gaacatcgcc tcgctccagt caatgaccgc tgttatgcgg ccattgtccg tcaggacatt
     5581 gttggagccg aaatccgcgt gcacgaggtg ccggacttcg gggcagtcct cggcccaaag
     5641 catcagctca tcgagagcct gcgcgacgga cgcactgacg gtgtcgtcca tcacagtttg
     5701 ccagtgatac acatggggat cagcaatcgc gcatatgaaa tcacgccatg tagtgtattg
     5761 accgattcct tgcggtccga atgggccgaa cccgctcgtc tggctaagat cggccgcagc
     5821 gatcgcatcc atggcctccg cgaccggctg cagaacagcg ggcagttcgg tttcaggcag
     5881 gtcttgcaac gtgacaccct gtgcacggcg ggagatgcaa taggtcaggc tctcgctgaa
     5941 ttccccaatg tcaagcactt ccggaatcgg gagcgcggcc gatgcaaagt gccgataaac
     6001 ataacgatct ttgtagaaac catcggcgca gctatttacc cgcaggacat atccacgccc
     6061 tcctacatcg aagctgaaag cacgagattc ttcgccctcc gagagctgca tcaggtcgga
     6121 gacgctgtcg aacttttcga tcagaaactt ctcgacagac gtcgcggtga gttcaggctt
     6181 tttcatggag cctgcttttt tgtacaaact tgcggcaccg cctgacatat taccttaaaa
     6241 ttcaaaaatt aatttcagat cagtcataac caagaaaaca aaataaacgt taggacagaa
     6301 aattcaaatt aattaaaaat tatgctgaat aagaaaatta catgcaaagc cttattttcc
     6361 tcatagaaac agttgaataa catatttttt aattgaaaac atagcaaaat tcgtggtaaa
     6421 atttgtcaaa accttggaaa aaatcgaaaa aagaacattt cccgatgcat tttcctattt
     6481 ctcgttgccg acaacgtata aacgttgaat actcagtcaa cgaatttggc tgctggcttg
     6541 gactctcgca cggaagagac gcgtgtgtag atttacgacg aaagcaaaaa tg
//
