LOCUS       sequence-193321-        5787 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  U626 Guide acceptor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5787)
  TITLE     Youles SynBio plasmids
REFERENCE   2  (bases 1 to 5787)
  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"
COMMENT     Sequence Label: pICSL1P5_AtU626
FEATURES             Location/Qualifiers
     source          1..5787
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(159..1307)
                     /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(1643..2231)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1723..1742)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2402..3193,3194..3262))
                     /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     3025..3044
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3263..3367)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3454..4085
                     /label=oriV
                     /note="incP origin of replication"
     misc_feature    4177..4201
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     protein_bind    4628..4649
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4664..4669,4670..4687,4688..4694)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4702..4718
                     /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)."
     RBS             4736..4747
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             4754..5431
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      5476..5547
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5563..5590
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_RNA        5604..5679
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(5691..5715)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    join(5766..5787,1..3)
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
ORIGIN
        1 aacctaagag aaaagagcgt ttattagaat aatcggatat ttaaaagggc gtgaaaaggt
       61 ttatccgttc gtccatttgt atgtgcatgc caaccacagg gttccccaga tcaggcgctg
      121 gctgctgaac ccccagccgg aactgacccc acaaggccct agcgtttgca atgcaccagg
      181 tcatcattga cccaggcgtg ttccaccagg ccgctgcctc gcaactcttc gcaggcttcg
      241 ccgacctgct cgcgccactt cttcacgcgg gtggaatccg atccgcacat gaggcggaag
      301 gtttccagct tgagcgggta cggctcccgg tgcgagctga aatagtcgaa catccgtcgg
      361 gccgtcggcg acagcttgcg gtacttctcc catatgaatt tcgtgtagtg gtcgccagca
      421 aacagcacga cgatttcctc gtcgatcagg acctggcaac gggacgtttt cttgccacgg
      481 tccaggacgc ggaagcggtg cagcagcgac accgattcca ggtgcccaac gcggtcggac
      541 gtgaagccca tcgccgtcgc ctgtaggcgc gacaggcatt cctcggcctt cgtgtaatac
      601 cggccattga tcgaccagcc caggtcctgg caaagctcgt agaacgtgaa ggtgatcggc
      661 tcgccgatag gggtgcgctt cgcgtactcc aacacctgct gccacaccag ttcgtcatcg
      721 tcggcccgca gctcgacgcc ggtgtaggtg atcttcacgt ccttgttgac gtggaaaatg
      781 accttgtttt gcagcgcctc gcgcgggatt ttcttgttgc gcgtggtgaa cagggcagag
      841 cgggccgtgt cgtttggcat cgctcgcatc gtgtccggcc acggcgcaat atcgaacaag
      901 gaaagctgca tttccttgat ctgctgcttc gtgtgtttca gcaacgcggc ctgcttggcc
      961 tcgctgacct gttttgccag gtcctcgccg gcggtttttc gcttcttggt cgtcatagtt
     1021 cctcgcgtgt cgatggtcat cgacttcgcc aaacctgccg cctcctgttc aagacgacgc
     1081 gaacgctcca cggcggccga tggcgcgggc agggcagggg gagccagttg cacgctgtcg
     1141 cgctcgatct tggccgtagc ttgctggacc atcgagccga cggactggaa ggtttcgcgg
     1201 ggcgcacgca tgacggtgcg gcttgcgatg gtttcggcat cctcggcgga aaaccccgcg
     1261 tcgatcagtt cttgcctgta tgccttccgg tcaaacgtcc gattcattca ccctccttgc
     1321 gggattgccc cgactcacgc cggggcaatg tgcccttatt cctgatttga cccgcctggt
     1381 gccttggtgt ccagataatc caccttatcg gcaatgaagt cggtcccgta gaccgtctgg
     1441 ccgtccttct cgtacttggt attccgaatc ttgccctgca cgaataccag cgaccccttg
     1501 cccaaatact tgccgtgggc ctcggcctga gagccaaaac acttgatgcg gaagaagtcg
     1561 gtgcgctcct gcttgtcgcc ggcatcgttg cgccacatct aggatctgcc aggaaccgta
     1621 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     1681 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     1741 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     1801 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     1861 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     1921 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     1981 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     2041 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     2101 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     2161 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     2221 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     2281 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     2341 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     2401 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     2461 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     2521 ccagtgctgc aatgataccg cgagaaccac gctcaccggc tccagattta tcagcaataa
     2581 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     2641 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     2701 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     2761 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     2821 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     2881 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     2941 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     3001 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     3061 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     3121 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     3181 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     3241 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcaag
     3301 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     3361 ttccgcgcac gaattggcca gcgctgccat ttttggggtg aggccgttcg cggccgaggg
     3421 gcgcagcccc tggggggatg ggaggcccgc gttagcgggc cgggagggtt cgagaagggg
     3481 gggcaccccc cttcggcgtg cgcggtcacg cgcacagggc gcagccctgg ttaaaaacaa
     3541 ggtttataaa tattggttta aaagcaggtt aaaagacagg ttagcggtgg ccgaaaaacg
     3601 ggcggaaacc cttgcaaatg ctggattttc tgcctgtgga cagcccctca aatgtcaata
     3661 ggtgcgcccc tcatctgtca gcactctgcc cctcaagtgt caaggatcgc gcccctcatc
     3721 tgtcagtagt cgcgcccctc aagtgtcaat accgcagggc acttatcccc aggcttgtcc
     3781 acatcatctg tgggaaactc gcgtaaaatc aggcgttttc gccgatttgc gaggctggcc
     3841 agctccacgt cgccggccga aatcgagcct gcccctcatc tgtcaacgcc gcgccgggtg
     3901 agtcggcccc tcaagtgtca acgtccgccc ctcatctgtc agtgagggcc aagttttccg
     3961 cgaggtatcc acaacgccgg cggccgcggt gtctcgcaca cggcttcgac ggcgtttctg
     4021 gcgcgtttgc agggccatag acggccgcca gcccagcggc gagggcaacc agcccggtga
     4081 gcgtcgcaaa ggagatcctg atctgactga tgggctgcct gtatcgagtg gtgattttgt
     4141 gccgagctgc cggtcgggga gctgttggct ggctggtggc aggatatatt gtggtgtaaa
     4201 caaattgacg cttagacaac ttaataacac attgcggacg tttttaatgt actggggtgg
     4261 atgcagtggg ccccactctg tgaagacaac agagaattcg catgcggagc atcttcattc
     4321 ttaagatatg aagataatct tcaaaaggcc cctgggaatc tgaaagaaga gaagcaggcc
     4381 catttatatg ggaaagaaca atagtatttc ttatataggc ccatttaagt tgaaaacaat
     4441 cttcaaaagt cccacatcgc ttagataaga aaacgaagct gagtttatat acagctagag
     4501 tcgaagtagt gattgtgaga cggatatcaa tacgcaaacc gcctctcccc gcgcgttggc
     4561 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     4621 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     4681 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacatac tagagaaaga
     4741 ggagaaatac tagatggctt cctccgagga cgttatcaaa gagttcatgc gtttcaaagt
     4801 tcgtatggaa ggttccgtta acggtcacga gttcgaaatc gaaggtgaag gtgaaggtcg
     4861 tccgtacgaa ggtacccaga ccgctaaact gaaagttacc aaaggtggtc cgctgccgtt
     4921 cgcttgggac atcctgtccc cgcagttcca gtacggttcc aaagcttacg ttaaacaccc
     4981 ggctgacatc ccggactacc tgaaactgtc cttcccggaa ggtttcaaat gggaacgtgt
     5041 tatgaacttc gaggacggtg gtgttgttac cgttacccag gactcctccc tgcaagacgg
     5101 tgagttcatc tacaaagtta aactgcgtgg taccaacttc ccgtccgacg gtccggttat
     5161 gcagaaaaaa accatgggtt gggaagcttc caccgaacgt atgtacccgg aggacggtgc
     5221 tctgaaaggt gaaatcaaaa tgcgtctgaa actgaaagac ggtggtcact acgacgctga
     5281 agttaaaacc acctacatgg ctaaaaaacc ggttcagctg ccgggtgctt acaaaaccga
     5341 catcaaactg gacatcacct cccacaacga ggactacacc atcgttgaac agtacgaacg
     5401 tgctgaaggt cgtcactcca ccggtgctta ataacgctga tagtgctagt gtagatcgct
     5461 actagagcca ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt
     5521 atctgttgtt tgtcggtgaa cgctctctac tagagtcaca ctggctcacc ttcgggtggg
     5581 cctttctgcg tttatacgtc tcagttttag agctagaaat agcaagttaa aataaggcta
     5641 gtccgttatc aacttgaaaa agtggcaccg agtcggtgct ttttttctag acccagcttt
     5701 cttgtacaaa gttggcatta cgcttgtgtt gtcttctgca cgaagtggtt taaactatca
     5761 gtgtttgaca ggatatattg gcgggta
//
