LOCUS       sequence-119208-        7225 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 1 Golden Gate Cassette: Plant hygromycin (with intron) 
            resistance cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7225)
  AUTHORS   Lawrenson T, Shorinola O, Stacey N, Li C, Ostergaard L, Patron N, 
            Uauy C, Harwood W
  TITLE     Induction of targeted, heritable mutations in barley and Brassica 
            oleracea using RNA-guided Cas9 nuclease.
  JOURNAL   Genome Biol. 2015 Nov 30;16(1):258. doi: 10.1186/s13059-015-0826-7.
  PUBMED    26616834
REFERENCE   2  (bases 1 to 7225)
  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: "Genome 
            Biol."; date: "2015-11-30"; volume: "16(1):258. doi"; pages: " 
            10.1186/s13059-015-0826-7"
COMMENT     Sequence Label: pICSL11059
FEATURES             Location/Qualifiers
     source          1..7225
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        994..1339
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     1282..1302
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     intron          1769..1958
                     /label=cat1 intron
                     /note="castor bean catalase intron, modified"
     polyA_signal    2680..2854
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     misc_feature    2899..2923
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(3079..4227)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     rep_origin      complement(4563..5151)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4643..4662)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5322..6113,6114..6182))
                     /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     5945..5964
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6183..6287)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6374..7005
                     /label=oriV
                     /note="incP origin of replication"
     misc_feature    7097..7121
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
ORIGIN
        1 ggaggaattc caatcccaca aaaatctgag cttaacagca cagttgctcc tctcagagca
       61 gaatcgggta ttcaacaccc tcatatcaac tactacgttg tgtataacgg tccacatgcc
      121 ggtatatacg atgactgggg ttgtacaaag gcggcaacaa acggcgttcc cggagttgca
      181 cacaagaaat ttgccactat tacagaggca agagcagcag ctgacgcgta cacaacaagt
      241 cagcaaacag acaggttgaa cttcatcccc aaaggagaag ctcaactcaa gcccaagagc
      301 tttgctaagg ccctaacaag cccaccaaag caaaaagccc actggctcac gctaggaacc
      361 aaaaggccca gcagtgatcc agccccaaaa gagatctcct ttgccccgga gattacaatg
      421 gacgatttcc tctatcttta cgatctagga aggaagttcg aaggtgaagg tgacgacact
      481 atgttcacca ctgataatga gaaggttagc ctcttcaatt tcagaaagaa tgctgaccca
      541 cagatggtta gagaggccta cgcagcaagt ctcatcaaga cgatctaccc gagtaacaat
      601 ctccaggaga tcaaatacct tcccaagaag gttaaagatg cagtcaaaag attcaggact
      661 aattgcatca agaacacaga gaaagacata tttctcaaga tcagaagtac tattccagta
      721 tggacgattc aaggcttgct tcataaacca aggcaagtaa tagagattgg agtctctaaa
      781 aaggtagttc ctactgaatc taaggccatg catggagtct aagattcaaa tcgaggatct
      841 aacagaactc gccgtcaaga ctggcgaaca gttcatacag agtcttttac gactcaatga
      901 caagaagaaa atcttcgtca acatggtgga gcacgacact ctggtctact ccaaaaatgt
      961 caaagataca gtctcagaag atcaaagggc tattgagact tttcaacaaa ggataatttc
     1021 gggaaacctc ctcggattcc attgcccagc tatctgtcac ttcatcgaaa ggacagtaga
     1081 aaaggaaggt ggctcctaca aatgccatca ttgcgataaa ggaaaggcta tcattcaaga
     1141 tctctctgcc gacagtggtc ccaaagatgg acccccaccc acgaggagca tcgtggaaaa
     1201 agaagaggtt ccaaccacgt ctacaaagca agtggattga tgtgacatct ccactgacgt
     1261 aagggatgac gcacaatccc actatccttc gcaagaccct tcctctatat aaggaagttc
     1321 atttcatttg gagaggacac gctcgagtat aagagctcat ttttacaaca attaccaaca
     1381 acaacaaaca acaaacaaca ttacaattac atttacaatt atcgatacaa tgaaaaagcc
     1441 tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga
     1501 cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg
     1561 tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta
     1621 tcggcacttt gcatcggccg cgctcccgat tccggaagtg cttgacattg gggaattcag
     1681 cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc
     1741 tgaaaccgaa ctgcccgctg ttctgcaggt aaatttctag tttttctcct tcattttctt
     1801 ggttaggacc cttttctctt tttatttttt tgagctttga tctttcttta aactgatcta
     1861 ttttttaatt gattggttat ggtgtaaata ttacatagct ttaactgata atctgattac
     1921 tttatttcgt gtgtctatga tgatgatgat aactgcagcc ggtcgcggag gccatggatg
     1981 cgatcgctgc ggccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa
     2041 tcggtcaata cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc
     2101 actggcaaac tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc
     2161 tgatgctttg ggccgaggac tgccccgaag tccggcacct cgtgcacgcg gatttcggct
     2221 ccaacaatgt cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga
     2281 tgttcgggga ttcccaatac gaggtcgcca acatcttctt ctggaggccg tggttggctt
     2341 gtatggagca gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc
     2401 ggctccgggc gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg
     2461 gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag
     2521 ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc ggccgtctgg accgatggct
     2581 gtgtagaagt actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg
     2641 aataggcttc tctagctaga gtcgatcgac aagctcgagt ttctccataa taatgtgtga
     2701 gtagttccca gataagggaa ttagggttcc tatagggttt cgctcatgtg ttgagcatat
     2761 aagaaaccct tagtatgtat ttgtatttgt aaaatacttc tatcaataaa atttctaatt
     2821 cctaaaacca aaatccagta ctaaaatcca gatcgctgca attgtcttct gcacgaagtg
     2881 gtttaaacta tcagtgtttg acaggatata ttggcgggta aacctaagag aaaagagcgt
     2941 ttattagaat aatcggatat ttaaaagggc gtgaaaaggt ttatccgttc gtccatttgt
     3001 atgtgcatgc caaccacagg gttccccaga tcaggcgctg gctgctgaac ccccagccgg
     3061 aactgacccc acaaggccct agcgtttgca atgcaccagg tcatcattga cccaggcgtg
     3121 ttccaccagg ccgctgcctc gcaactcttc gcaggcttcg ccgacctgct cgcgccactt
     3181 cttcacgcgg gtggaatccg atccgcacat gaggcggaag gtttccagct tgagcgggta
     3241 cggctcccgg tgcgagctga aatagtcgaa catccgtcgg gccgtcggcg acagcttgcg
     3301 gtacttctcc catatgaatt tcgtgtagtg gtcgccagca aacagcacga cgatttcctc
     3361 gtcgatcagg acctggcaac gggacgtttt cttgccacgg tccaggacgc ggaagcggtg
     3421 cagcagcgac accgattcca ggtgcccaac gcggtcggac gtgaagccca tcgccgtcgc
     3481 ctgtaggcgc gacaggcatt cctcggcctt cgtgtaatac cggccattga tcgaccagcc
     3541 caggtcctgg caaagctcgt agaacgtgaa ggtgatcggc tcgccgatag gggtgcgctt
     3601 cgcgtactcc aacacctgct gccacaccag ttcgtcatcg tcggcccgca gctcgacgcc
     3661 ggtgtaggtg atcttcacgt ccttgttgac gtggaaaatg accttgtttt gcagcgcctc
     3721 gcgcgggatt ttcttgttgc gcgtggtgaa cagggcagag cgggccgtgt cgtttggcat
     3781 cgctcgcatc gtgtccggcc acggcgcaat atcgaacaag gaaagctgca tttccttgat
     3841 ctgctgcttc gtgtgtttca gcaacgcggc ctgcttggcc tcgctgacct gttttgccag
     3901 gtcctcgccg gcggtttttc gcttcttggt cgtcatagtt cctcgcgtgt cgatggtcat
     3961 cgacttcgcc aaacctgccg cctcctgttc aagacgacgc gaacgctcca cggcggccga
     4021 tggcgcgggc agggcagggg gagccagttg cacgctgtcg cgctcgatct tggccgtagc
     4081 ttgctggacc atcgagccga cggactggaa ggtttcgcgg ggcgcacgca tgacggtgcg
     4141 gcttgcgatg gtttcggcat cctcggcgga aaaccccgcg tcgatcagtt cttgcctgta
     4201 tgccttccgg tcaaacgtcc gattcattca ccctccttgc gggattgccc cgactcacgc
     4261 cggggcaatg tgcccttatt cctgatttga cccgcctggt gccttggtgt ccagataatc
     4321 caccttatcg gcaatgaagt cggtcccgta gaccgtctgg ccgtccttct cgtacttggt
     4381 attccgaatc ttgccctgca cgaataccag cgaccccttg cccaaatact tgccgtgggc
     4441 ctcggcctga gagccaaaac acttgatgcg gaagaagtcg gtgcgctcct gcttgtcgcc
     4501 ggcatcgttg cgccacatct aggatctgcc aggaaccgta aaaaggccgc gttgctggcg
     4561 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4621 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4681 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4741 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4801 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4861 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4921 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4981 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     5041 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     5101 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     5161 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     5221 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     5281 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     5341 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     5401 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     5461 cgagaaccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     5521 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5581 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5641 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5701 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5761 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5821 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5881 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5941 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     6001 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     6061 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     6121 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     6181 atactcttcc tttttcaata ttattgaagc atttatcaag gttattgtct catgagcgga
     6241 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac gaattggcca
     6301 gcgctgccat ttttggggtg aggccgttcg cggccgaggg gcgcagcccc tggggggatg
     6361 ggaggcccgc gttagcgggc cgggagggtt cgagaagggg gggcaccccc cttcggcgtg
     6421 cgcggtcacg cgcacagggc gcagccctgg ttaaaaacaa ggtttataaa tattggttta
     6481 aaagcaggtt aaaagacagg ttagcggtgg ccgaaaaacg ggcggaaacc cttgcaaatg
     6541 ctggattttc tgcctgtgga cagcccctca aatgtcaata ggtgcgcccc tcatctgtca
     6601 gcactctgcc cctcaagtgt caaggatcgc gcccctcatc tgtcagtagt cgcgcccctc
     6661 aagtgtcaat accgcagggc acttatcccc aggcttgtcc acatcatctg tgggaaactc
     6721 gcgtaaaatc aggcgttttc gccgatttgc gaggctggcc agctccacgt cgccggccga
     6781 aatcgagcct gcccctcatc tgtcaacgcc gcgccgggtg agtcggcccc tcaagtgtca
     6841 acgtccgccc ctcatctgtc agtgagggcc aagttttccg cgaggtatcc acaacgccgg
     6901 cggccgcggt gtctcgcaca cggcttcgac ggcgtttctg gcgcgtttgc agggccatag
     6961 acggccgcca gcccagcggc gagggcaacc agcccggtga gcgtcgcaaa ggagatcctg
     7021 atctgactga tgggctgcct gtatcgagtg gtgattttgt gccgagctgc cggtcgggga
     7081 gctgttggct ggctggtggc aggatatatt gtggtgtaaa caaattgacg cttagacaac
     7141 ttaataacac attgcggacg tttttaatgt actggggtgg atgcagtggg ccccactctg
     7201 tgaagacaat gccgaattcg gatcc
//
