LOCUS       sequence-337471-        9157 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  cargo vector for the phiC31 system with Psur5:GFP, cb_unc-119, and 
            Gateway R1R2 site (for gene of interest insertion).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9157)
  AUTHORS   Yang FJ, Chen CN, Chang T, Cheng TW, Chang NC, Kao CY, Lee CC, Huang
            YC, Hsu JC, Li J, Lu MJ, Chan SP, Wang J
  TITLE     phiC31 integrase for recombination-mediated single-copy insertion 
            and genome manipulation in Caenorhabditis elegans.
  JOURNAL   Genetics. 2022 Feb 4;220(2):iyab206. doi: 10.1093/genetics/iyab206.
  PUBMED    34791215
REFERENCE   2  (bases 1 to 9157)
  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: "Genetics.";
            date: "2022-02-4"; volume: "220(2):iyab206. doi"; pages: " 
            10.1093/genetics/iyab206"
COMMENT     Sequence Label: pBRC_double_attB_GFP_donor
FEATURES             Location/Qualifiers
     source          1..9157
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    28..61
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     primer_bind     complement(1230..1258)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2006..2026
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             complement(2242..2253)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(4741..5046)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(4843..4862)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(5388..6047)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5962..5981
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(6101..6107,6108..6125,6126..6131))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(6156..6280)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    6530..6563
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     primer_bind     complement(6607..6629)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6729..6748
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6826..6844)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6912..7016
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7017..7085,7086..7877)
                     /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(7235..7254)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8048..8636
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8537..8556
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8790..8807
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8924..8945
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8960..8965,8966..8983,8984..8990)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8998..9014
                     /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     9003..9025
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     9022..9038
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     9022..9038
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 gaaatgaaat aagcttgggc tgcacgggtg ccagggcgtg cccttgggct ccccgggcgc
       61 gtactgcaga gtggagagtg tagtccctcg tcggctctct gtgtatttct ttcaatgcct
      121 gtgtctcggt ctatctttgt ctctatctgt tttaaaagtt ttttgcaaag tttcaatttc
      181 ttttttattt tctcttcttt ttttgcacat ttattttttt taaagagcgc tgttcttcaa
      241 ttggaaaaca gtataacgat cgaaataaaa aaaaaacttg aaatagtatc ggaattgaga
      301 tgggggcaat tgaaattgaa gacgcaacaa aaacatgttt ttatcgtctg aaattatcta
      361 aactttgaat ctaacgtgtc aatggctact tttacagtag actttttaga gtaatgttaa
      421 aaaggataat ataggttttt cttcaaacta ttttgtcacg ccgcatatat tcttttcata
      481 ttccattcgc aaagtgcatg atgtaacaaa aatgaaacgt cataattttt taatctttca
      541 cacttttttt gccggccgtt aaatcaaaat caaatcaact taatccaaca cgtgcgggct
      601 tgcaatttta ttacaccggt gtcctacagc gccatgttat cttctttgat tagagatctc
      661 tcacgcaaca tttgacattg aaaattttgt ctccgaattg ttgtgtacct actttgtccc
      721 gatgtcccaa attttcactg ataacaagag atatctcaac tgttcttatc acgttgatac
      781 gtagaacgtt ttacactaaa ccaaattgca gtccccttct acgaaccgtt tgctaattgt
      841 tttgcataga acagtgatca atgtgttgtg ctcactagtc aatgatgttc gtctctcatt
      901 tctcctacaa cttcactgaa aaggttttga atatctggtg attgtaggca atcacaccac
      961 agttaatggc ctggcaaata atgataagcg tgtgataaac ctcacaaaaa tacgcctaga
     1021 agtggtgata tcgctagttt tgtgtccttt cgattttttt cgtgttgctt ttttctgtgg
     1081 ataaaccttt gaactttaca ttttgttttc agaataaccg ctctagagga tcctatcgat
     1141 tcgcggccgc tgtacacccg ggtgccctat agtgagtcgt attggtaccg gtagaaaaaa
     1201 tgagtaaagg agaagaactt ttcactggag ttgtcccaat tcttgttgaa ttagatggtg
     1261 atgttaatgg gcacaaattt tctgtcagtg gagagggtga aggtgatgca acatacggaa
     1321 aacttaccct taaatttatt tgcactactg gaaaactacc tgttccatgg gtaagtttaa
     1381 acatatatat actaactaac cctgattatt taaattttca gccaacactt gtcactactt
     1441 tctgttatgg tgttcaatgc ttctcgagat acccagatca tatgaaacag catgactttt
     1501 tcaagagtgc catgcccgaa ggttatgtac aggaaagaac tatatttttc aaagatgacg
     1561 ggaactacaa gacacgtaag tttaaacagt tcggtactaa ctaaccatac atatttaaat
     1621 tttcaggtgc tgaagtcaag tttgaaggtg atacccttgt taatagaatc gagttaaaag
     1681 gtattgattt taaagaagat ggaaacattc ttggacacaa attggaatac aactataact
     1741 cacacaatgt atacatcatg gcagacaaac aaaagaatgg aatcaaagtt gtaagtttaa
     1801 acatgatttt actaactaac taatctgatt taaattttca gaacttcaaa attagacaca
     1861 acattgaaga tggaagcgtt caactagcag accattatca acaaaatact ccaattggcg
     1921 atggccctgt ccttttacca gacaaccatt acctgtccac acaatctgcc ctttcgaaag
     1981 atcccaacga aaagagagac cacatggtcc ttcttgagtt tgtaacagct gctgggatta
     2041 cacatggcat ggacgaacta tacaaaagcc cacaagcttt ggaattcgct agcctttgag
     2101 ccaatttatc caagtccttg taaaaagtat tcgaaaattg ttaacggata aaatgtttat
     2161 tataatatca aaaacaattt gtcagttgac cactttttga tatatttgac agaaaccggg
     2221 atgaattggc tcaaagttag ggcgcccttc tattacaggg tttctgataa caaaccggtt
     2281 attaactccc aacaagggat gatttcaatt catcatgctc aaattgaccc aaattaagtt
     2341 acatgacaaa ttcatcgccc ttttcaactc ttcttggctc atcatctgtt attctgttcc
     2401 tattctgtct gcaccctata cccctttgca tactctctcg tcattccttt cttggatagt
     2461 gtcttcttct ccccagctct gctacttcta tgacttgcgc ccgctgcttt tccgctcgtt
     2521 cctctcctct ctgacgtctt cttctgcttc ttttctcctt cctttttccc ttctcaatct
     2581 tcctcttttt cccatttctg tcaatcattc gaagaagaag aagaagaaga ccctcattca
     2641 tttcattttt ttttctgtcg gtgtgtgctg ctcggttaag agtgagctct ctattccacg
     2701 tcttcttctt tttcttcttc gattcgaatc aatcactcca ccaaaacgca ttcgtttttg
     2761 ggattcaccc cgcggttcgc aataggtttc tttttcaaat tattagcgtt ataaatagaa
     2821 aaatggtgga gtttcaataa aaatgataat ttacaaagtg tattttgatt acatgtactc
     2881 aaaaaggttg aaatttttca taccagtttt ccggaaatcc atctgatatc attatcgtat
     2941 ttttcttttt aaaaatgttt tcaaaaaaaa acaaaatata gctggttatt tggcaccctc
     3001 taattaccat tttcctgtca caccacactc ttttccttct tccacttctt tccgttttca
     3061 gccgcttcca accaaaccga tatgaaagcc gagcaacaac gatcgattcc acccggctcg
     3121 gcgaccttcc cgtcgcaggt gagactcaga aaactagaga aaccgctcaa ctaactcttg
     3181 atatccgatt tcattctttt ctcttttctt tttgttgaac ttttccatat ttccagatgc
     3241 cacggccacc accaagcacc gaacaaggaa tcacaacgga atcggagctt gcgaagaaag
     3301 ctcaaatcac tccgaacgac gttttagcac ttccgggaat cactcaaggt atcttgctcc
     3361 ctttggattt cagtttaaaa cataaattta ggattcttat gctccccatc tgcgaacatc
     3421 tataacatcg agttcaccaa gttccaaatc cgtgatctgg acactgagca agtgcttttc
     3481 gagatcgcca aaccggagaa cgatcaggag aatgatgagt cgccacagga gtcggcaaga
     3541 tacgtgcgtt atagatttgc tccaaacttt ttgaaactca aaacggtcgg agcaactgtg
     3601 gaattcaaag taggagacat cccaatccat catttccgaa tgatcgaacg tcacttcttc
     3661 aaagatcgcc ttctgaagtg ttttgacttt gaattcggat tctgtattcc gaattcacga
     3721 aacaactgtg aacatatcta tgagttccct caactctctc aacaactcag tgagtcatta
     3781 ttctaaaaag tacaattcaa aagactaatc tctttttcag tggacgacat gatcaacaat
     3841 ccaaacgaga ctcgttctga cagcttctat ttcgtcgata acaaactcgt catgcacaac
     3901 aaagccgact actcatatga tgcataaata tttaatacaa aaatgttctg gataattatt
     3961 ctgtcgaata gaaaaaaaac tccaaatgtg attaaattcc aataattcct gtctagtttg
     4021 ttccttcccc ttccccttct catgttcaat gcattcctaa gcttttcagt tccccccttg
     4081 tttctatatt ttttcgctgt cctgtcacac tcgctaaaaa cactaatcac acggaaatct
     4141 gttttcaata aaaactccaa ctttaactca ttttcaattt caactgaaag attttttcat
     4201 tagagaatgt cgctagcctt tgagccaatt tatcccggtt tctgtcaaat atatcaaaaa
     4261 gtggtcaact gacaaattgt ttttgatatt ataataaaca ttttatccgt taacaatttt
     4321 cgaatacttt ttacaaggac ttggataaat tggctcaaag gctagccggc catacaagta
     4381 atccggatga tcgacgccaa cgtcgttgaa ttttcaaatt ttaaatactg aatatttgtt
     4441 ttttttccta ttatttattt attctctttg tgtttttttt cttgctttct aaaaaattaa
     4501 ttcaatccaa atctaaacat ttttttttct ctttccgtct cccaattcgt attccgctcc
     4561 tctcatctga acacgtgcac tttgtacaag aaagctgaac gagaaacgta aaatgatata
     4621 aatatcaata tattaaatta gattttgcat aaaaaacaga ctacataata ctgtaaaaca
     4681 caacatatcc agtcactatg gtcgacctgc agactggctg tgtataaggg agcctgacat
     4741 ttatattccc cagaacatca ggttaatggc gtttttgatg tcattttcgc ggtggctgag
     4801 atcagccact tcttccccga taacggagac cggcacactg gccatatcgg tggtcatcat
     4861 gcgccagctt tcatccccga tatgcaccac cgggtaaagt tcacgggaga ctttatctga
     4921 cagcagacgt gcactggcca gggggatcac catccgtcgc ccgggcgtgt caataatatc
     4981 actctgtaca tccacaaaca gacgataacg gctctctctt ttataggtgt aaaccttaaa
     5041 ctgcatttca ccagcccctg ttctcgtcag caaaagagcc gttcatttca ataaaccggg
     5101 cgacctcagc catcccttcc tgattttccg ctttccagcg ttcggcacgc agacgacggg
     5161 cttcattctg catggttgtg cttaccagac cggagatatt gacatcatat atgccttgag
     5221 caactgatag ctgtcgctgt caactgtcac tgtaatacgc tgcttcatag catacctctt
     5281 tttgacatac ttcgggtata catatcagta tatattctta taccgcaaaa atcagcgcgc
     5341 aaatacgcat actgttatct ggcttttagt aagccggatc cagatcttta cgccccgccc
     5401 tgccactcat cgcagtactg ttgtaattca ttaagcattc tgccgacatg gaagccatca
     5461 caaacggcat gatgaacctg aatcgccagc ggcatcagca ccttgtcgcc ttgcgtataa
     5521 tatttgccca tggtgaaaac gggggcgaag aagttgtcca tattggccac gtttaaatca
     5581 aaactggtga aactcaccca gggattggct gagacgaaaa acatattctc aataaaccct
     5641 ttagggaaat aggccaggtt ttcaccgtaa cacgccacat cttgcgaata tatgtgtaga
     5701 aactgccgga aatcgtcgtg gtattcactc cagagcgatg aaaacgtttc agtttgctca
     5761 tggaaaacgg tgtaacaagg gtgaacacta tcccatatca ccagctcacc gtctttcatt
     5821 gccatacgga attccggatg agcattcatc aggcgggcaa gaatgtgaat aaaggccgga
     5881 taaaacttgt gcttattttt ctttacggtc tttaaaaagg ccgtaatatc cagctgaacg
     5941 gtctggttat aggtacattg agcaactgac tgaaatgcct caaaatgttc tttacgatgc
     6001 cattgggata tatcaacggt ggtatatcca gtgatttttt tctccatttt agcttcctta
     6061 gctcctgaaa atctcgacgg atcctaactc aaaatccaca cattatacga gccggaagca
     6121 taaagtgtaa agcctggggt gcctaatgcg gccgccatag tgactggata tgttgtgttt
     6181 tacagtatta tgtagtctgt tttttatgca aaatctaatt taatatattg atatttatat
     6241 cattttacgt ttctcgttca gcttttttgt acaaacttgt cacgtgcaag tttatttatc
     6301 ttctcgcttt catttcatta ggacgtgggg ggaattggtg gaagggggaa acacacaaaa
     6361 ggatgatgga aatgaaataa ggacacacaa tatgcaacaa cattcaattc agaaatatgg
     6421 aggaaggttt aaaagaaaac ataaaaatat atagaggagg aaggaaaact agtaaaaaat
     6481 aagcaaagaa attaggcgaa cgatgagaat tgtcctcgct tgggccgtac gcgcccgggg
     6541 agcccaaggg cacgccctgg cacccgggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     6601 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     6661 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     6721 taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc
     6781 gcacagatgc gtaaggagaa aataccgcat caggcggcct taagggcctc gtgatacgcc
     6841 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     6901 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     6961 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     7021 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     7081 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     7141 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     7201 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     7261 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     7321 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     7381 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     7441 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     7501 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     7561 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     7621 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     7681 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     7741 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     7801 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     7861 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     7921 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     7981 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     8041 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     8101 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     8161 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     8221 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     8281 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     8341 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     8401 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     8461 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     8521 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     8581 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     8641 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     8701 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     8761 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     8821 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     8881 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca
     8941 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg
     9001 tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc aagctgtaag
     9061 tttaaacatg atcttactaa ctaactattc tcatttaaat tttcagagct taaaaatggc
     9121 tgaaatcact cacaacgatg gatacgctaa caacttg
//
