LOCUS       sequence-496035-        6039 bp    DNA     circular SYN 02-JAN-2026
DEFINITION  Level 1 Position 2 FORWARD acceptor Low - copy (with pBR322 origin 
            of replication) GGAG - GCTT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6039)
  TITLE     Plant Golden-gate Promoters, Terminators and Level0/Level1 cloning 
            acceptors.
REFERENCE   2  (bases 1 to 6039)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 2, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pICSL47742-LC
FEATURES             Location/Qualifiers
     source          1..6039
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    96..117
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(132..137,138..155,156..162)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    170..186
                     /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     175..197
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     194..210
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     194..210
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             206..529
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    223..279
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(280..297)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(280..296)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(289..311)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     misc_feature    671..695
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(851..1999)
                     /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      2465..3053
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2954..2973
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3207..3224
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3239..3379
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3465..3487
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3481..3672)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(4165..4956,4957..5025))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDECDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4788..4807
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5026..5130)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5217..5848
                     /label=oriV
                     /note="incP origin of replication"
     misc_feature    5940..5964
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
ORIGIN
        1 tctgtgaaga caagcaagaa ttcaagcttg gagtgagacc gcagctggca cgacaggttt
       61 cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag
      121 gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga
      181 taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc atgcctgcag
      241 gtcgactcta gaggatcccc gggtaccgag ctcgaattca ctggccgtcg ttttacaacg
      301 tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt
      361 cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
      421 cctgaatggc gaatggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
      481 acaccgcata tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
      541 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
      601 ttacagacaa gctgtgacgg tctcacgcta ctattgtctt ctgcacgaag tggtttaaac
      661 tatcagtgtt tgacaggata tattggcggg taaacctaag agaaaagagc gtttattaga
      721 ataatcggat atttaaaagg gcgtgaaaag gtttatccgt tcgtccattt gtatgtgcat
      781 gccaaccaca gggttcccca gatcaggcgc tggctgctga acccccagcc ggaactgacc
      841 ccacaaggcc ctagcgtttg caatgcacca ggtcatcatt gacccaggcg tgttccacca
      901 ggccgctgcc tcgcaactct tcgcaggctt cgccgacctg ctcgcgccac ttcttcacgc
      961 gggtggaatc cgatccgcac atgaggcgga aggtttccag cttgagcggg tacggctccc
     1021 ggtgcgagct gaaatagtcg aacatccgtc gggccgtcgg cgacagcttg cggtacttct
     1081 cccatatgaa tttcgtgtag tggtcgccag caaacagcac gacgatttcc tcgtcgatca
     1141 ggacctggca acgggacgtt ttcttgccac ggtccaggac gcggaagcgg tgcagcagcg
     1201 acaccgattc caggtgccca acgcggtcgg acgtgaagcc catcgccgtc gcctgtaggc
     1261 gcgacaggca ttcctcggcc ttcgtgtaat accggccatt gatcgaccag cccaggtcct
     1321 ggcaaagctc gtagaacgtg aaggtgatcg gctcgccgat aggggtgcgc ttcgcgtact
     1381 ccaacacctg ctgccacacc agttcgtcat cgtcggcccg cagctcgacg ccggtgtagg
     1441 tgatcttcac gtccttgttg acgtggaaaa tgaccttgtt ttgcagcgcc tcgcgcggga
     1501 ttttcttgtt gcgcgtggtg aacagggcag agcgggccgt gtcgtttggc atcgctcgca
     1561 tcgtgtccgg ccacggcgca atatcgaaca aggaaagctg catttccttg atctgctgct
     1621 tcgtgtgttt cagcaacgcg gcctgcttgg cctcgctgac ctgttttgcc aggtcctcgc
     1681 cggcggtttt tcgcttcttg gtcgtcatag ttcctcgcgt gtcgatggtc atcgacttcg
     1741 ccaaacctgc cgcctcctgt tcaagacgac gcgaacgctc cacggcggcc gatggcgcgg
     1801 gcagggcagg gggagccagt tgcacgctgt cgcgctcgat cttggccgta gcttgctgga
     1861 ccatcgagcc gacggactgg aaggtttcgc ggggcgcacg catgacggtg cggcttgcga
     1921 tggtttcggc atcctcggcg gaaaaccccg cgtcgatcag ttcttgcctg tatgccttcc
     1981 ggtcaaacgt ccgattcatt caccctcctt gcgggattgc cccgactcac gccggggcaa
     2041 tgtgccctta ttcctgattt gacccgcctg gtgccttggt gtccagataa tccaccttat
     2101 cggcaatgaa gtcggtcccg tagaccgtct ggccgtcctt ctcgtacttg gtattccgaa
     2161 tcttgccctg cacgaatacc agcgacccct tgcccaaata cttgccgtgg gcctcggcct
     2221 gagagccaaa acacttgatg cggaagaagt cggtgcgctc ctgcttgtcg ccggcatcgt
     2281 tgcgccacat ctaagatcca gaccaagttt actcatatat actttagatt gatttaaaac
     2341 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     2401 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     2461 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     2521 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     2581 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     2641 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     2701 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     2761 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     2821 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     2881 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     2941 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3001 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     3061 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     3121 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     3181 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     3241 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc
     3301 tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg
     3361 tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     3421 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gactccggga gctgcatgtg
     3481 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc
     3541 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt
     3601 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc
     3661 ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc
     3721 gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact
     3781 ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac
     3841 tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca
     3901 gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag
     3961 actttacgaa acacggaaac ccaagaccat tcatgttgtt gctcaggtcg cagacgtttt
     4021 gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag
     4081 gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg
     4141 ccaggaccca acgctgcccg agccttacca atgcttaatc agtgaggcac ctatctcagc
     4201 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     4261 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagaac cacgctcacc
     4321 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     4381 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     4441 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcaca
     4501 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     4561 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     4621 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     4681 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     4741 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     4801 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     4861 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     4921 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     4981 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     5041 atattattga agcatttatc aaggttattg tctcatgagc ggatacatat ttgaatgtat
     5101 ttagaaaaat aaacaaatag gggttccgcg cacgaattgg ccagcgctgc catttttggg
     5161 gtgaggccgt tcgcggccga ggggcgcagc ccctgggggg atgggaggcc cgcgttagcg
     5221 ggccgggagg gttcgagaag ggggggcacc ccccttcggc gtgcgcggtc acgcgcacag
     5281 ggcgcagccc tggttaaaaa caaggtttat aaatattggt ttaaaagcag gttaaaagac
     5341 aggttagcgg tggccgaaaa acgggcggaa acccttgcaa atgctggatt ttctgcctgt
     5401 ggacagcccc tcaaatgtca ataggtgcgc ccctcatctg tcagcactct gcccctcaag
     5461 tgtcaaggat cgcgcccctc atctgtcagt agtcgcgccc ctcaagtgtc aataccgcag
     5521 ggcacttatc cccaggcttg tccacatcat ctgtgggaaa ctcgcgtaaa atcaggcgtt
     5581 ttcgccgatt tgcgaggctg gccagctcca cgtcgccggc cgaaatcgag cctgcccctc
     5641 atctgtcaac gccgcgccgg gtgagtcggc ccctcaagtg tcaacgtccg cccctcatct
     5701 gtcagtgagg gccaagtttt ccgcgaggta tccacaacgc cggcggccgc ggtgtctcgc
     5761 acacggcttc gacggcgttt ctggcgcgtt tgcagggcca tagacggccg ccagcccagc
     5821 ggcgagggca accagcccgg tgagcgtcgc aaaggagatc ctgatctgac tgatgggctg
     5881 cctgtatcga gtggtgattt tgtgccgagc tgccggtcgg ggagctgttg gctggctggt
     5941 ggcaggatat attgtggtgt aaacaaattg acgcttagac aacttaataa cacattgcgg
     6001 acgtttttaa tgtactgggg tggatgcagt gggccccac
//
