LOCUS       sequence-447542-        3167 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 2 Destination vector - Position B - Assembly of 4-5 TU's from 
            Level 1-ABCD or ABCDE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3167)
  AUTHORS   Moore SJ, Lai HE, Kelwick RJ, Chee SM, Bell DJ, Polizzi KM, Freemont
            PS
  TITLE     EcoFlex: A Multifunctional MoClo Kit for E. coli Synthetic Biology.
  JOURNAL   ACS Synth Biol. 2016 May 2.
  PUBMED    27096716
REFERENCE   2  (bases 1 to 3167)
  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: "ACS Synth 
            Biol. 2016 May 2."
COMMENT     Sequence Label: pTU2-B-RFP
FEATURES             Location/Qualifiers
     source          1..3167
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    123..144
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(159..164,165..182,183..189)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    197..213
                     /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             231..242
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             249..926
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      971..1042
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1058..1085
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      1120..1177
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(1201..1218)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1372..1960)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1452..1471)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2143..2237)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2258..2917)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2832..2851
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     3079..3097
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(3101..3144)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    join(3147..3167,1)
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
ORIGIN
        1 ggtctcagta catctagaga cgcaatacgc aaaccgcctc tccccgcgcg ttggccgatt
       61 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
      121 attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct
      181 cgtatgttgt gtggaattgt gagcggataa caatttcaca catactagag aaagaggaga
      241 aatactagat ggcttcctcc gaagacgtta tcaaagagtt catgcgtttc aaagttcgta
      301 tggaaggttc cgttaacggt cacgagttcg aaatcgaagg tgaaggtgaa ggtcgtccgt
      361 acgaaggtac ccagaccgct aaactgaaag ttaccaaagg tggtccgctg ccgttcgctt
      421 gggacatcct gtccccgcag ttccagtacg gttccaaagc ttacgttaaa cacccggctg
      481 acatcccgga ctacctgaaa ctgtccttcc cggaaggttt caaatgggaa cgtgttatga
      541 acttcgaaga cggtggtgtt gttaccgtta cccaggactc ctccctgcaa gacggtgagt
      601 tcatctacaa agttaaactg cgtggtacca acttcccgtc cgacggtccg gttatgcaga
      661 aaaaaaccat gggttgggaa gcttccaccg aacgtatgta cccggaagac ggtgctctga
      721 aaggtgaaat caaaatgcgt ctgaaactga aagacggtgg tcactacgac gctgaagtta
      781 aaaccaccta catggctaaa aaaccggttc agctgccggg tgcttacaaa accgacatca
      841 aactggacat cacctcccac aacgaagact acaccatcgt tgaacagtac gaacgtgctg
      901 aaggtcgtca ctccaccggt gcttaataac gctgatagtg ctagtgtaga tcgctactag
      961 agccaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
     1021 ttgtttgtcg gtgaacgctc tctactagag tcacactggc tcaccttcgg gtgggccttt
     1081 ctgcgtttat acgtctcatt agggacagag accctgcagt ccggcaaaaa agggcaaggt
     1141 gtcaccaccc tgcccttttt ctttaaaacc gaaaagatta cttcgcgtta tgcaggcttc
     1201 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     1261 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     1321 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccacag
     1381 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     1441 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1501 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1561 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     1621 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     1681 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     1741 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     1801 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     1861 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     1921 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     1981 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     2041 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     2101 aagtatatat gagtaaactt ggtctgacag ctcgaggctt ggattctcac caataaaaaa
     2161 cgcccggcgg caaccgagcg ttctgaacaa atccagatgg agttctgagg tcattactgg
     2221 atctatcaac aggagtccaa gcgagctcga tatcaaatta cgccccgccc tgccactcat
     2281 cgcagtactg ttgtaattca ttaagcattc tgccgacatg gaagccatca caaacggcat
     2341 gatgaacctg aatcgccagc ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca
     2401 tggtgaaaac gggggcgaag aagttgtcca tattggccac gtttaaatca aaactggtga
     2461 aactcaccca gggattggct gacacgaaaa acatattctc aataaaccct ttagggaaat
     2521 aggccaggtt ttcaccgtaa cacgccacat cttgcgaata tatgtgtaga aactgccgga
     2581 aatcgtcgtg gtattcactc cagagcgatg aaaacgtttc agtttgctca tggaaaacgg
     2641 tgtaacaagg gtgaacacta tcccatatca ccagctcacc gtctttcatt gccatacgaa
     2701 attccggatg agcattcatc aggcgggcaa gaatgtgaat aaaggccgga taaaacttgt
     2761 gcttattttt ctttacggtc tttaaaaagg ccgtaatatc cagctgaacg gtctggttat
     2821 aggtacattg agcaactgac tgaaatgcct caaaatgttc tttacgatgc cattgggata
     2881 tatcaacggt ggtatatcca gtgatttttt tctccatttt agcttcctta gctcctgaaa
     2941 atctcgataa ctcaaaaaat acgcccggta gtgatcttat ttcattatgg tgaaagttgg
     3001 aacctcttac gtgcccgatc aactcgagtg ccacctgacg tctaagaaac cattattatc
     3061 atgacattaa cctataaaaa taggcgtatc acgaggcaga atttcagata aaaaaaatcc
     3121 ttagctttcg ctaaggatga tttctggaat tcgcggccgc ttctaga
//
