LOCUS       sequence-390622-        4436 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor vector to restore L1 destination via Esp3I (reverse 
            orientation).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4436)
  AUTHORS   Chamness JC, Kumar J, Cruz AJ, Rhuby E, Holum MJ, Cody JP, Tibebu R,
            Gamo ME, Starker CG, Zhang F, Voytas DF
  TITLE     An extensible vector toolkit and parts library for advanced 
            engineering of plant genomes.
  JOURNAL   Plant Genome. 2023 Mar 9:e20312. doi: 10.1002/tpg2.20312.
  PUBMED    36896468
REFERENCE   2  (bases 1 to 4436)
  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: "Plant 
            Genome. 2023 Mar 9:e20312. doi: 10.1002/tpg2.20312."
COMMENT     Sequence Label: pJMC-L1-Reverse-Donor-Esp3I
FEATURES             Location/Qualifiers
     source          1..4436
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     68..86
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     392..411
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        392..410
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        join(506..511,512..529,530..536)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             590..1270
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     CDS             1590..1895
                     /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     1774..1793
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     promoter        complement(join(2355..2361,2362..2379,2380..2385))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    2400..2421
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2541..2558)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2712..3300)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2792..2811)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3471..4262,4263..4331))
                     /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     4094..4113
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4332..4436)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
       61 ctataaaaat aggcgtatca cgaggccgcc cctgcagccg aattatatta tttttgccaa
      121 ataattttta acaaaagctc tgaagtcttc ttcatttaaa ttcttagatg atacttcatc
      181 tggaaaattg tcccaattag tagcatcacg ctgtgagtaa gttctaaacc atttttttat
      241 tgttgtatta tctctaatct tactactcga tgagttttcg gtattatctc tatttttaac
      301 ttggagcagg ttccattcat tgtttttttc atcatagtga ataaaatcaa ctgctttaac
      361 acttgtgcct gaacaccata tccatccggc gtaatacgac tcactatagg gagagcggcc
      421 gccagatctt ccggatggct cgagtttttc agcaagattt tcgtctctac cgagcgtgag
      481 accggccgca ttaggcaccc caggctttac actttatgct tccggctcgt ataatgtgtg
      541 gattttgagt taggatccgt cgagattttc aggagctaag gaagctaaaa tggagaaaaa
      601 aatcactgga tataccaccg ttgatatatc ccaatggcat cgtaaagaac attttgaggc
      661 atttcagtca gtcgctcaat gtacctataa ccagaccgtt cagctggata ttacggcctt
      721 tttaaagacc gtaaagaaaa ataagcacaa gttttatccg gcctttattc acattcttgc
      781 ccgcctgatg aatgctcatc cggaattccg tatggcaatg aaagacggtg agctggtgat
      841 atgggatagt gttcaccctt gttacaccgt tttccatgag caaactgaaa cgttttcatc
      901 gctctggagt gaataccacg acgatttccg gcagtttcta cacatatatt cgcaagatgt
      961 ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg tttattgaga atatgttttt
     1021 cgtttcagcc aatccctggg tgagtttcac cagttttgat ttaaacgtgg ccaatatgga
     1081 caacttcttc gcccccgttt tcaccatggg caaatattat acgcaaggcg acaaggtgct
     1141 gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat ggcttccatg tcggcagaat
     1201 gcttaatgaa ttacaacagt actgcgatga gtggcaggcg gggcgtaatc tagaggatcc
     1261 ggcttactaa aagccagata acagtatgcg tatttgcgcg ctgatttttg cggtataaga
     1321 atatatactg atatgtatac ccgaagtatg tcaaaaggag gtatgctatg aagcagcgta
     1381 ttacagtgac agttgacagc gacagctatc agttgctcaa ggcatatatg atgtcaatat
     1441 ctccggtctg gtaagcacaa ccatgcagaa tgaagcccgt cgtctgcgtg ccgaacgctg
     1501 gaaagcggaa aatcaggaag ggatggctga ggtcgcccgg tttattgaaa tgaacggctc
     1561 ttttgctgac gagaacaggg gctggtgaaa tgcagtttaa ggtttacacc tataaaagag
     1621 agagccgtta tcgtctgttt gtggatgtac agagtgatat tattgacacg cccgggcgac
     1681 ggatggtgat ccccctggcc agtgcacgtc tgctgtcaga taaagtctcc cgtgaacttt
     1741 acccggtggt gcatatcggg gatgaaagct ggcgcatgat gaccaccgat atggccagtg
     1801 tgccggtttc cgttatcggg gaagaagtgg ctgatctcag ccaccgcgaa aatgacatca
     1861 aaaacgccat taacctgatg ttctggggaa tataaggtct cactccagtg agagacgaaa
     1921 atctttctag aagatctcct acaatattct cagctgccat ggaaaatcga tgttcttctt
     1981 ttattctctc aagattttca ggctgtatat taaaacttat attaagaact atgctaacca
     2041 cctcatcagg aaccgttgta ggtggcgtgg gttttcttgg caatcgactc tcatgaaaac
     2101 tacgagctaa atattcaata tgttcctctt gaccaacttt attctgcatt ttttttgaac
     2161 gaggtttaga gcaagcttca ggaaactgag acaggaattt tattaaaaat ttaaattttg
     2221 aagaaagttc agggttaata gcatccattt tttgctttgc aagttcctca gcattcttaa
     2281 caaaagatgt ctcttttgac atgtttaaag tttaaacctc ctgtgtgaaa ttattatccg
     2341 ctcataattc cacacattat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     2401 tgagtgagct aactcacatt aattgcgttg cgctcactgc caattgcttt ccagtcggga
     2461 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     2521 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     2581 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     2641 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     2701 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     2761 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     2821 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     2881 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     2941 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     3001 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     3061 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     3121 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     3181 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     3241 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     3301 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     3361 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     3421 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     3481 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     3541 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     3601 atgataccgc gtgacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     3661 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     3721 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     3781 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     3841 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     3901 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     3961 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     4021 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     4081 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     4141 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     4201 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     4261 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     4321 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     4381 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcg
//
