LOCUS       sequence-266105-        6165 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a recombineering cassette for tagging a target gene within a
            genomic clone with a fluorescent protein. Cassette: FRT-galK-FRT in 
            the 2nd intron of gfp.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6165)
  AUTHORS   Tursun B, Cochella L, Carrera I, Hobert O
  TITLE     A toolkit and robust pipeline for the generation of fosmid-based 
            reporter genes in C. elegans.
  JOURNAL   PLoS One. 2009;4(3):e4625. doi: 10.1371/journal.pone.0004625. Epub 
            2009 Mar 4.
  PUBMED    19259264
REFERENCE   2  (bases 1 to 6165)
  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: "PLoS One.";
            date: "2009"; volume: "4(3):e4625. doi"; pages: " 
            10.1371/journal.pone.0004625. Epub 2009 Mar 4"
COMMENT     Sequence Label: pBALU1
FEATURES             Location/Qualifiers
     source          1..6165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     17..39
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     31..48
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     32..48
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     55..74
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        55..73
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(174..202)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     protein_bind    complement(542..575)
                     /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)"
     promoter        579..626
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     primer_bind     complement(1808..1824)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1808..1824)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    complement(1833..1866)
                     /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)"
     primer_bind     2275..2295
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     promoter        complement(2415..2433)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2416..2433)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2451..2467)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2451..2467)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2464..2486)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2475..2491
                     /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)."
     promoter        complement(join(2499..2505,2506..2523,2524..2529))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2544..2565
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2682..2699)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2853..3441)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2933..2952)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3612..4403,4404..4472))
                     /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     4235..4254
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             complement(4490..5284)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4601..4620)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5211..5230
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      complement(5628..6056)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5738..5759)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5950..5969
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga attgtaatac
       61 gactcactat agggcgaatt gggccctcta gatgcatgct cgagcggccg ccagtgtgat
      121 ggatatctgc agaattcgcc cttatgagta aaggagaaga acttttcact ggagttgtcc
      181 caattcttgt tgaattagat ggtgatgtta atgggcacaa attttctgtc agtggagagg
      241 gtgaaggtga tgcaacatac ggaaaactta cccttaaatt tatttgcact actggaaaac
      301 tacctgttcc atgggtaagt ttaaacatat atatactaac taaccctgat tatttaaatt
      361 ttcagccaac acttgtcact actttctgtt atggtgttca atgcttctcg agatacccag
      421 atcatatgaa acggcatgac tttttcaaga gtgccatgcc cgaaggttat gtacaggaaa
      481 gaactatatt tttcaaagat gacgggaact acaagacacg taagtttaaa cagttcggta
      541 cgaagttcct atactttcta gagaatagga acttccctgt tgacaattaa tcatcggcat
      601 agtatatcgg catagtataa tacgacaagg tgaggaacta aacccaggag gcagatcatg
      661 agtctgaaag aaaaaacaca atctctgttt gccaacgcat ttggctaccc tgccactcac
      721 accattcagg cgcctggccg cgtgaatttg attggtgaac acaccgacta caacgacggt
      781 ttcgttctgc cctgcgcgat tgattatcaa accgtgatca gttgtgcacc acgcgatgac
      841 cgtaaagttc gcgtgatggc agccgattat gaaaatcagc tcgacgagtt ttccctcgat
      901 gcgcccattg tcgcacatga aaactatcaa tgggctaact acgttcgtgg cgtggtgaaa
      961 catctgcaac tgcgtaacaa cagcttcggc ggcgtggaca tggtgatcag cggcaatgtg
     1021 ccgcagggtg ccgggttaag ttcttccgct tcactggaag tcgcggtcgg aaccgtattg
     1081 cagcagcttt atcatctgcc gctggacggc gcacaaatcg cgcttaacgg tcaggaagca
     1141 gaaaaccagt ttgtaggctg taactgcggg atcatggatc agctaatttc cgcgctcggc
     1201 aagaaagatc atgccttgct gatcgattgc cgctcactgg ggaccaaagc agtttccatg
     1261 cccaaaggtg tggctgtcgt catcatcaac agtaacttca aacgtaccct ggttggcagc
     1321 gaatacaaca cccgtcgtga acagtgcgaa accggtgcgc gtttcttcca gcagccagcc
     1381 ctgcgtgatg tcaccattga agagttcaac gctgttgcgc atgaactgga cccgatcgtg
     1441 gcaaaacgcg tgcgtcatat actgactgaa aacgcccgca ccgttgaagc tgccagcgcg
     1501 ctggagcaag gcgacctgaa acgtatgggc gagttgatgg cggagtctca tgcctctatg
     1561 cgcgatgatt tcgaaatcac cgtgccgcaa attgacactc tggtagaaat cgtcaaagct
     1621 gtgattggcg acaaaggtgg cgtacgcatg accggcggcg gatttggcgg ctgtatcgtc
     1681 gcgctgatcc cggaagagct ggtgcctgcc gtacagcaag ctgtcgctga acaatatgaa
     1741 gcaaaaacag gtattaaaga gactttttac gtttgtaaac catcacaagg agcaggacag
     1801 tgctgaggat ccactagttc tagagcggcc gcgaagttcc tatactttct agagaatagg
     1861 aacttctaac taaccataca tatttaaatt ttcaggtgct gaagtcaagt ttgaaggtga
     1921 tacccttgtt aatagaatcg agttaaaagg tattgatttt aaagaagatg gaaacattct
     1981 tggacacaaa ttggaataca actataactc acacaatgta tacatcatgg cagacaaaca
     2041 aaagaatgga atcaaagttg taagtttaaa catgatttta ctaactaact aatctgattt
     2101 aaattttcag aacttcaaaa ttagacacaa cattgaagat ggaagcgttc aactagcaga
     2161 ccattatcaa caaaatactc caattggcga tggccctgtc cttttaccag acaaccatta
     2221 cctgtccaca caatctgccc tttcgaaaga tcccaacgaa aagagagacc acatggtcct
     2281 tcttgagttt gtaacagctg ctgggattac acatggcatg gatgaactat acaaaaaggg
     2341 cgaattccag cacactggcg gccgttacta gtggatccga gctcggtacc aagcttgatg
     2401 catagcttga gtattctata gtgtcaccta aatagcttgg cgtaatcatg gtcatagctg
     2461 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     2521 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     2581 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     2641 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     2701 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     2761 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     2821 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     2881 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     2941 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     3001 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     3061 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     3121 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     3181 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     3241 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     3301 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     3361 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     3421 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     3481 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     3541 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     3601 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     3661 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     3721 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     3781 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     3841 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     3901 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     3961 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     4021 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     4081 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     4141 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     4201 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     4261 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     4321 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     4381 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     4441 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaatt cagaagaact
     4501 cgtcaagaag gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca
     4561 cgaggaagcg gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg
     4621 ctatgtcctg atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc
     4681 ggccattttc caccatgata ttcggcaagc aggcatcgcc atgggtcacg acgagatcct
     4741 cgccgtcggg catgcgcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat
     4801 gctcttcgtc cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct
     4861 cgatgcgatg tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc
     4921 gccgcattgc atcagccatg atggatactt tctcggcagg agcaaggtgg gatgacagga
     4981 gatcctgccc cggcacttcg cccaatagca gccagtccct tcccgcttca gtgacaacgt
     5041 cgagcacagc tgcgcaagga acgcccgtcg tggccagcca cgatagccgc gctgcctcgt
     5101 cctgcagttc attcagggca ccggacaggt cggtcttgac aaaaagaacc gggcgcccct
     5161 gcgctgacag ccggaacacg gcggcatcag agcagccgat tgtctgttgt gcccagtcat
     5221 agccgaatag cctctccacc caagcggccg gagaacctgc gtgcaatcca tcttgttcaa
     5281 tcatgcgaaa cgatcctcat cctgtctctt gatcagatct tgatcccctg cgccatcaga
     5341 tccttggcgg caagaaagcc atccagttta ctttgcaggg cttcccaacc ttaccagagg
     5401 gcgccccagc tggcaattcc ggttcgcttg ctgtccataa aaccgcccag tctagctatc
     5461 gccatgtaag cccactgcaa gctacctgct ttctctttgc gcttgcgttt tcccttgtcc
     5521 agatagccca gtagctgaca ttcatccggg gtcagcaccg tttctgcgga ctggctttct
     5581 acgtgttccg cttcctttag cagcccttgc gccctgaatt ttgttaaaat tcgcgttaaa
     5641 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa
     5701 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact
     5761 attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc
     5821 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     5881 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     5941 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     6001 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccat
     6061 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
     6121 cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgg
//
