LOCUS       sequence-486015-        5039 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Hyg resistant attB phiC31 recombination-mediation insertion (RMI) 
            vector for C. elegans.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5039)
  AUTHORS   Beck AA, Nonet ML
  TITLE     A single component landing site for efficient transgenesis using 
            recombination-mediated insertion.
  JOURNAL   MicroPubl Biol. 2025 Sep 2;2025:10.17912/micropub.biology.001784. 
            doi: 10.17912/micropub.biology.001784. eCollection 2025.
  PUBMED    40970081
REFERENCE   2  (bases 1 to 5039)
  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: "MicroPubl 
            Biol."; date: "2025-09-2"; volume: 
            "2025:10.17912/micropub.biology.001784. doi"; pages: " 
            10.17912/micropub.biology.001784. eCollection 2025"
COMMENT     Sequence Label: pAttBF3Hyg
FEATURES             Location/Qualifiers
     source          1..5039
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    15..48
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     protein_bind    112..145
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    520..544
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(545..1570)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(545..1570)
                     /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(1573..1597)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    1996..2029
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(2134..2167)
                     /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)"
     primer_bind     complement(2205..2224)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2206..2224)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2234..2251)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2234..2250)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2243..2265)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2392..2847
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2479..2498)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2689..2710
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2873..2977
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2978..3046,3047..3838)
                     /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     complement(3196..3215)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4009..4597
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4498..4517
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4751..4768
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4884..4905
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4920..4925,4926..4943,4944..4950)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4958..4974
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     4963..4985
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4982..4998
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4982..4998
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5017..5037
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5019..5037
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 aagccgcggt gcgggtgcca gggcgtgccc ttgggctccc cgggcgcgta ctccacctca
       61 cccatctggt ccatcatgat gaacgggtcg aggtggcggt agttgataca aataacttcg
      121 tataaagtat cctatacgaa gttatcccag ttgtcctcga acaggtctaa aaattgcggt
      181 cataaactga aacgtaacat atgataaggt attttgtgtg cggttttttc tatgatgttt
      241 caaaaaaatt tatcagaagt aaaaaacata agaagtgtac acagaacatt gtgtagattt
      301 tgggatagaa ttattattaa tggggggagg gagcacaatt tttttcgact ttttattacg
      361 aattaattct atttttgaat ctacacaatt tcattgttag aggtgactta aaagaagcta
      421 aaaaacaaag aaattaagag aagttttttg ataataacaa aaataggggg tgggagcaca
      481 gggagaaaga gcatgtaggg atgttgaaga gtaattggac ccactttgta caagaaagct
      541 gggtctattc ctttgccctc ggacgagtgc tggggcgtcg gtttccacta tcggcgagta
      601 cttctacaca gccatcggtc cagacggccg cgcttctgcg ggcgatttgt gtacgcccga
      661 cagtcccggc tccggatcgg acgattgcgt cgcatcgacc ctgcgcccaa gctgcatcat
      721 cgaaattgcc gtcaaccaag ctctgataga gttggtcaag accaatgcgg agcatatacg
      781 cccggagccg cggcgatcct gcaagctccg gatgcctccg ctcgaagtag cgcgtctgct
      841 gctccataca agccaaccac ggcctccaga agaagatgtt ggcgacctcg tattgggaat
      901 ccccgaacat cgcctcgctc cagtcaatga ccgctgttat gcggccattg tccgtcagga
      961 cattgttgga gccgaaatcc gcgtgcacga ggtgccggac ttcggggcag tcctcggccc
     1021 aaagcatcag ctcatcgaga gcctgcgcga cggacgcact gacggtgtcg tccatcacag
     1081 tttgccagtg atacacatgg ggatcagcaa tcgcgcatat gaaatcacgc catgtagtgt
     1141 attgaccgat tccttgcggt ccgaatgggc cgaacccgct cgtctggcta agatcggccg
     1201 cagcgatcgc atccatggcc tccgcgaccg gctgcagaac agcgggcagt tcggtttcag
     1261 gcaggtcttg caacgtgaca ccctgtgcac ggcgggagat gcaataggtc aggctctcgc
     1321 tgaattcccc aatgtcaagc acttccggaa tcgggagcgc ggccgatgca aagtgccgat
     1381 aaacataacg atctttgtag aaaccatcgg cgcagctatt tacccgcagg acatatccac
     1441 gccctcctac atcgaagctg aaagcacgag attcttcgcc ctccgagagc tgcatcaggt
     1501 cggagacgct gtcgaacttt tcgatcagaa acttctcgac agacgtcgcg gtgagttcag
     1561 gctttttcat ggagcctgct tttttgtaca aacttgcggc accgcctgac atattacctt
     1621 aaaattcaaa aattaatttc agatcagtca taaccaagaa aacaaaataa acgttaggac
     1681 agaaaattca aattaattaa aaattatgct gaataagaaa attacatgca aagccttatt
     1741 ttcctcatag aaacagttga ataacatatt ttttaattga aaacatagca aaattcgtgg
     1801 taaaatttgt caaaaccttg gaaaaaatcg aaaaaagaac atttcccgat gcattttcct
     1861 atttctcgtt gccgacaacg tataaacgtt gaatactcag tcaacgaatt tggctgctgg
     1921 cttggactct cgcacggaag agacgcgtgt gtagatttac gacgaaagca aaaatgatcc
     1981 cgctttccag tcgggataac ttcgtataaa gtatcctata cgaagttata agctggagct
     2041 ccaccgcggt ggcggccgct ctagatggcg aagagcccat ggatccacta gtaacatatg
     2101 tgcaggcctg ctcttcggta agcttatcga tacgaagttc ctatactatt tgaagaatag
     2161 gaacttccgt cgacccgtta accgggggcc cggtacccaa ttcgccctat agtgagtcgt
     2221 attacgcgcg ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     2281 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     2341 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg gacgcgccct
     2401 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     2461 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     2521 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     2581 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     2641 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     2701 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     2761 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     2821 ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac
     2881 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     2941 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     3001 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     3061 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     3121 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     3181 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     3241 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     3301 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     3361 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     3421 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     3481 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     3541 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     3601 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     3661 tattgctgat aaatctggag ccggtgagcg tggatctcgc ggtatcattg cagcactggg
     3721 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     3781 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     3841 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     3901 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     3961 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     4021 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     4081 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     4141 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     4201 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     4261 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     4321 gggctgaatg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     4381 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     4441 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     4501 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     4561 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     4621 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     4681 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
     4741 gaccgagcgc agcgagtcag tgagcgagga agcggaattc gcccaatacg caaaccgcct
     4801 ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa
     4861 gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct
     4921 ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata acaatttcac
     4981 acaggaaaca gctatgacca tgattacgcc aagcgcgcaa ttaaccctca ctaaaggga
//
