LOCUS       sequence-443110-        9000 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plant Transformation Vector that allows marker removal through 
            Mendelian inheritance. Start of DNA transfer at RB, terminates at 
            left border but in ~1/3 of cases goes ahead until second left 
            border.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9000)
  TITLE     Gene Tracking
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 9000)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pCambia-dLB-kanR-GFP
FEATURES             Location/Qualifiers
     source          1..9000
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    398..422
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     misc_feature    573..597
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     protein_bind    complement(848..881)
                     /label=lox66
                     /bound_moiety="Cre recombinase"
                     /note="Right element (RE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1067..1412
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     1355..1375
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     regulatory      1429..1438
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2164..2217
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     terminator      3028..3279
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     protein_bind    complement(3284..3317)
                     /label=lox71
                     /bound_moiety="Cre recombinase"
                     /note="Left element (LE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3340..3362
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
     CDS             3568..4362
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     rep_origin      4449..5037
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4938..4957
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5191..5208
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5223..5363
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5289..5308)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5449..5471
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     rep_origin      5707..5901
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     CDS             complement(5967..7040)
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     CDS             complement(7469..8098)
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
ORIGIN
        1 tcggcccgtt ggttggtcaa gtcctggtcg tcggtgctga cgcgggcata gcccagcagg
       61 ccagcggcgg cgctcttgtt catggcgtaa tgtctccggt tctagtcgca agtattctac
      121 tttatgcgac taaaacacgc gacaagaaaa cgccaggaaa agggcagggc ggcagcctgt
      181 cgcgtaactt aggacttgtg cgacatgtcg ttttcagaag acggctgcac tgaacgtcag
      241 aagccgactg cactatagca gcggaggggt tggatcaaag tactttgatc ccgaggggaa
      301 ccctgtggtt ggcatgcaca tacaaatgga cgaacggata aaccttttca cgccctttta
      361 aatatccgtt attctaataa acgctctttt ctcttaggtt tacccgccaa tatatcctgt
      421 caaacactga tagtttaaac ctaggccttg cactagtaaa caaaagcttc caacatggtg
      481 gaattcacaa agctacctta agagagtcta gagttaattc agtacattaa aaacgtccgc
      541 aatgtgttat taagttgtct aagcgtcaat ttgtttacac cacaatatat cctgccacca
      601 gccagccaac agctccccga ccggcagctc ggcacaaaat caccactcga tacaggcagc
      661 ccatcagtcc gggacggcgt cagcgggaga gccgttgtaa ggcggcagac tttgctcatg
      721 ttaccgatgc tattcggaag aacggcaact aagctgccgg gtttgaaaca cggatgatct
      781 cgcggagggt agcatgttga ttgtaacgat gacagagcgt tgctgcctgt gatcagtcga
      841 caaattcata acttcgtata gcatacatta tacgaacggt atgcatggag tcaaagattc
      901 aaatagagga cctaacagaa ctcgccgtaa agactggcga acagttcata cagagtctct
      961 tacgactcaa tgacaagaag aaaatcttcg tcaacatggt ggagcacgac acacttgtct
     1021 actccaaaaa tatcaaagat acagtctcag aagaccaaag ggcaattgag acttttcaac
     1081 aaagggtaat atccggaaac ctcctcggat tccattgccc agctatctgt cactttattg
     1141 tgaagatagt ggaaaaggaa ggtggctcct acaaatgcca tcattgcgat aaaggaaagg
     1201 ccatcgttga agatgcctct gccgacagtg gtcccaaaga tggaccccca cccacgagga
     1261 gcatcgtgga aaaagaagac gttccaacca cgtcttcaaa gcaagtggat tgatgtgata
     1321 tctccactga cgtaagggat gacgcacaat cccactatcc ttcgcaagac ccttcctcta
     1381 tataaggaag ttcatttcat ttggagagaa cacggtggac tcttgactgc caccatggtt
     1441 tcatctggag aagatatatt ctcaggatta gtcccaattc taatagaact ggagggtgat
     1501 gttaatggac acagattcag tgtcagaggt gaaggatatg gtgatgcctc caatgggaaa
     1561 ctggaaatta aatttatatg taccactgga agattacctg taccttggcc tactcttgta
     1621 accaccttat cctatggtgt acagtgcttt gcaaagtatc ctgagcacat gaggcagaat
     1681 gacttcttca aatctgccat gccagatggt tatgtccaag agaggactat cagtttcaaa
     1741 gaggatggaa cctacaagac cagagctgaa gtcaagttcg agggtgaagc tctggtaaac
     1801 aggattgacc tcaagggcct tgaatttaaa gaggatggaa acattcttgg tcataaacta
     1861 gaatatagtt tcaacagcca ctacgtgtat attactgctg acaagaatag gaatggtctg
     1921 gaggctcagt ttagaattag acacaatgta gatgatggct ctgtccaact tgctgaccat
     1981 taccaacaga atacacctat aggtgagggt cctgtcctac ttcctgagca acattatctc
     2041 actaccaata gtgtcctaag taaggaccct caagagagaa gagatcacat ggtgctggtt
     2101 gaatttgtga ctgctgctgg cctgtcacta ggaatggatg aactctacaa gagtggatct
     2161 ggtgagggca gaggaagtct tctaacatgt ggtgatgtgg aggagaatcc tggtcccatg
     2221 attgaacaag atggattgca tgcaggttct cctgctgctt gggtggagag gctatttggc
     2281 tatgactggg cacaacagac aataggctgc tctgatgctg cagtgttcag actgtcagct
     2341 cagggtagac ctgttctctt tgtcaagact gacctgtctg gtgccctgaa tgaactccaa
     2401 gatgaggcag ctagactatc ttggctggcc actacaggtg ttccttgtgc agctgtgctt
     2461 gatgttgtca ctgaagctgg aagggactgg ctgctattgg gtgaagtgcc tgggcaggat
     2521 ctcctgtcat ctcaccttgc tcctgctgag aaagtatcca tcatggctga tgcaatgaga
     2581 agactgcata ctcttgatcc tgctacctgc ccattcgacc accaagctaa acatagaatt
     2641 gagagaggaa gaactagaat ggaagctggt cttgtggatc aggatgatct ggatgaagag
     2701 catcagggtc ttgctccagc agaactgttt gccaggctca aggctagaat gcctgatggt
     2761 gaggatcttg tggtgaccca gggtgatgcc tgcttgccta atatcatggt ggaaaatggc
     2821 agattctctg gattcattga ctgtggcaga ctgggtgtgg ctgacagata tcaggacata
     2881 gctttggcta ccagagatat tgctgaagag cttggtggag aatgggctga cagattcctt
     2941 gtgctttatg gtattgctgc tcctgattct cagagaattg ccttctatag acttcttgat
     3001 gagttcttct aagactcttt cgaagcagat cgttcaaaca tttggcaata aagtttctta
     3061 agattgaatc ctgttgccgg tcttgcgatg attatcatat aatttctgtt gaattacgtt
     3121 aagcatgtaa taattaacat gtaatgcatg acgttattta tgagatgggt tttttgatta
     3181 gagtcccgca attatacatt taatacgcga tagaaaacaa aatatagcgc gcaaactagg
     3241 ataaattatc gcgcgcggtg tcatctatgt tactagatcg atctaccgtt cgtatagcat
     3301 acattatacg aagttatacc ggcatgcaag ctgataacgt tacaccacaa tatatcctgc
     3361 caagatctct aattccgcgg ctcgcggttt caaaatcggc tccgtcgata ctatgttata
     3421 cgccaacttt gaaaacaact ttgaaaaagc tgttttctgg tatttaaggt tttagaatgc
     3481 aaggaacagt gaattggagt tcgtcttgtt ataattagct tcttggggta tctttaaata
     3541 ctgtagaaaa gaggaaggaa ataataaatg gctaaaatga gaatatcacc ggaattgaaa
     3601 aaactgatcg aaaaataccg ctgcgtaaaa gatacggaag gaatgtctcc tgctaaggta
     3661 tataagctgg tgggagaaaa tgaaaaccta tatttaaaaa tgacggacag ccggtataaa
     3721 gggaccacct atgatgtgga acgggaaaag gacatgatgc tatggctgga aggaaagctg
     3781 cctgttccaa aggtcctgca ctttgaacgg catgatggct ggagcaatct gctcatgagt
     3841 gaggccgatg gcgtcctttg ctcggaagag tatgaagatg aacaaagccc tgaaaagatt
     3901 atcgagctgt atgcggagtg catcaggctc tttcactcca tcgacatatc ggattgtccc
     3961 tatacgaata gcttagacag ccgcttagcc gaattggatt acttactgaa taacgatctg
     4021 gccgatgtgg attgcgaaaa ctgggaagaa gacactccat ttaaagatcc gcgcgagctg
     4081 tatgattttt taaagacgga aaagcccgaa gaggaacttg tcttttccca cggcgacctg
     4141 ggagacagca acatctttgt gaaagatggc aaagtaagtg gctttattga tcttgggaga
     4201 agcggcaggg cggacaagtg gtatgacatt gccttctgcg tccggtcgat cagggaggat
     4261 atcggggaag aacagtatgt cgagctattt tttgacttac tggggatcaa gcctgattgg
     4321 gagaaaataa aatattatat tttactggat gaattgtttt agtacctaga atgcatgacc
     4381 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     4441 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     4501 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     4561 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     4621 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     4681 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     4741 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     4801 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     4861 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     4921 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     4981 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     5041 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     5101 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     5161 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag
     5221 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatggtgc
     5281 actctcagta caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc
     5341 tacgtgactg ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac
     5401 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     5461 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag gcagggtgcc ttgatgtggg
     5521 cgccggcggt cgagtggcga cggcgcggct tgtccgcgcc ctggtagatt gcctggccgt
     5581 aggccagcca tttttgagcg gccagcggcc gcgataggcc gacgcgaagc ggcggggcgt
     5641 agggagcgca gcgaccgaag ggtaggcgct ttttgcagct cttcggctgt gcgctggcca
     5701 gacagttatg cacaggccag gcgggtttta agagttttaa taagttttaa agagttttag
     5761 gcggaaaaat cgcctttttt ctcttttata tcagtcactt acatgtgtga ccggttccca
     5821 atgtacggct ttgggttccc aatgtacggg ttccggttcc caatgtacgg ctttgggttc
     5881 ccaatgtacg tgctatccac aggaaagaga ccttttcgac ctttttcccc tgctagggca
     5941 atttgcccta gcatctgctc cgtacattag gaaccggcgg atgcttcgcc ctcgatcagg
     6001 ttgcggtagc gcatgactag gatcgggcca gcctgccccg cctcctcctt caaatcgtac
     6061 tccggcaggt catttgaccc gatcagcttg cgcacggtga aacagaactt cttgaactct
     6121 ccggcgctgc cactgcgttc gtagatcgtc ttgaacaacc atctggcttc tgccttgcct
     6181 gcggcgcggc gtgccaggcg gtagagaaaa cggccgatgc cgggatcgat caaaaagtaa
     6241 tcggggtgaa ccgtcagcac gtccgggttc ttgccttctg tgatctcgcg gtacatccaa
     6301 tcagctagct cgatctcgat gtactccggc cgcccggttt cgctctttac gatcttgtag
     6361 cggctaatca aggcttcacc ctcggatacc gtcaccaggc ggccgttctt ggccttcttc
     6421 gtacgctgca tggcaacgtg cgtggtgttt aaccgaatgc aggtttctac caggtcgtct
     6481 ttctgctttc cgccatcggc tcgccggcag aacttgagta cgtccgcaac gtgtggacgg
     6541 aacacgcggc cgggcttgtc tcccttccct tcccggtatc ggttcatgga ttcggttaga
     6601 tgggaaaccg ccatcagtac caggtcgtaa tcccacacac tggccatgcc ggccggccct
     6661 gcggaaacct ctacgtgccc gtctggaagc tcgtagcgga tcacctcgcc agctcgtcgg
     6721 tcacgcttcg acagacggaa aacggccacg tccatgatgc tgcgactatc gcgggtgccc
     6781 acgtcataga gcatcggaac gaaaaaatct ggttgctcgt cgcccttggg cggcttccta
     6841 atcgacggcg caccggctgc cggcggttgc cgggattctt tgcggattcg atcagcggcc
     6901 gcttgccacg attcaccggg gcgtgcttct gcctcgatgc gttgccgctg ggcggcctgc
     6961 gcggccttca acttctccac caggtcatca cccagcgccg cgccgatttg taccgggccg
     7021 gatggtttgc gaccgctcac gccgattcct cgggcttggg ggttccagtg ccattgcagg
     7081 gccggcaggc aacccagccg cttacgcctg gccaaccgcc cgttcctcca cacatggggc
     7141 attccacggc gtcggtgcct ggttgttctt gattttccat gccgcctcct ttagccgcta
     7201 aaattcatct actcatttat tcatttgctc atttactctg gtagctgcgc gatgtattca
     7261 gatagcagct cggtaatggt cttgccttgg cgtaccgcgt acatcttcag cttggtgtga
     7321 tcctccgccg gcaactgaaa gttgacccgc ttcatggctg gcgtgtctgc caggctggcc
     7381 aacgttgcag ccttgctgct gcgtgcgctc ggacggccgg cacttagcgt gtttgtgctt
     7441 ttgctcattt tctctttacc tcattaactc aaatgagttt tgatttaatt tcagcggcca
     7501 gcgcctggac ctcgcgggca gcgtcgccct cgggttctga ttcaagaacg gttgtgccgg
     7561 cggcggcagt gcctgggtag ctcacgcgct gcgtgatacg ggactcaaga atgggcagct
     7621 cgtacccggc cagcgcctcg gcaacctcac cgccgatgcg cgtgcctttg atcgcccgcg
     7681 acacgacaaa ggccgcttgt agccttccat ccgtgacctc aatgcgctgc ttaaccagct
     7741 ccaccaggtc ggcggtggcc catatgtcgt aagggcttgg ctgcaccgga atcagcacga
     7801 agtcggctgc cttgatcgcg gacacagcca agtccgccgc ctggggcgct ccgtcgatca
     7861 ctacgaagtc gcgccggccg atggccttca cgtcgcggtc aatcgtcggg cggtcgatgc
     7921 cgacaacggt tagcggttga tcttcccgca cggccgccca atcgcgggca ctgccctggg
     7981 gatcggaatc gactaacaga acatcggccc cggcgagttg cagggcgcgg gctagatggg
     8041 ttgcgatggt cgtcttgcct gacccgcctt tctggttaag tacagcgata accttcatgc
     8101 gttccccttg cgtatttgtt tatttactca tcgcatcata tacgcagcga ccgcatgacg
     8161 caagctgttt tactcaaata cacatcacct ttttagacgg cggcgctcgg tttcttcagc
     8221 ggccaagctg gccggccagg ccgccagctt ggcatcagac aaaccggcca ggatttcatg
     8281 cagccgcacg gttgagacgt gcgcgggcgg ctcgaacacg tacccggccg cgatcatctc
     8341 cgcctcgatc tcttcggtaa tgaaaaacgg ttcgtcctgg ccgtcctggt gcggtttcat
     8401 gcttgttcct cttggcgttc attctcggcg gccgccaggg cgtcggcctc ggtcaatgcg
     8461 tcctcacgga aggcaccgcg ccgcctggcc tcggtgggcg tcacttcctc gctgcgctca
     8521 agtgcgcggt acagggtcga gcgatgcacg ccaagcagtg cagccgcctc tttcacggtg
     8581 cggccttcct ggtcgatcag ctcgcgggcg tgcgcgatct gtgccggggt gagggtaggg
     8641 cgggggccaa acttcacgcc tcgggccttg gcggcctcgc gcccgctccg ggtgcggtcg
     8701 atgattaggg aacgctcgaa ctcggcaatg ccggcgaaca cggtcaacac catgcggccg
     8761 gccggcgtgg tggtgtcggc ccacggctct gccaggctac gcaggcccgc gccggcctcc
     8821 tggatgcgct cggcaatgtc cagtaggtcg cgggtgctgc gggccaggcg gtctagcctg
     8881 gtcactgtca caacgtcgcc agggcgtagg tggtcaagca tcctggccag ctccgggcgg
     8941 tcgcgcctgg tgccggtgat cttctcggaa aacagcttgg tgcagccggc cgcgtgcagt
//
