LOCUS       40924_33967        6130 bp DNA     circular SYN 18-DEC-2018
DEFINITION  MISSA donor vector pP-ccdB, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6130)
  AUTHORS   Chen QJ, Xie M, Ma XX, Dong L, Chen J, Wang XC.
  TITLE     MISSA is a highly efficient in vivo DNA assembly method for plant 
            multiple-gene transformation
  JOURNAL   Plant Physiol. 153 (1), 41-51 (2010)
  PUBMED    20200068
REFERENCE   2  (bases 1 to 6130)
  AUTHORS   Chen Q-J., Xie M, Ma X-X., Dong L, Chen J, Wang X-C.
  TITLE     Direct Submission
  JOURNAL   Submitted (27-JAN-2010) State Key Laboratory of Plant Physiology and
            Biochemistry, College of Biological Sciences, China Agricultural 
            University, No. 2 West Yuanmingyuan Road, Haidian, Beijing 100193, 
            China
REFERENCE   3  (bases 1 to 6130)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6130)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Plant 
            Physiol."; date: "2010"; volume: "153"; issue: "1"; pages: "41-51"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (27-JAN-2010) State Key Laboratory of Plant Physiology and 
            Biochemistry, College of Biological Sciences, China Agricultural 
            University, No. 2 West Yuanmingyuan Road, Haidian, Beijing 100193, 
            China"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6130
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(1..34)
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      100..186
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      278..305
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     primer_bind     373..389
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        397..415
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     protein_bind    424..655
                     /label=attP1
                     /note="recombination site for the Gateway(R) BP reaction"
     terminator      complement(733..760)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(852..938)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     rep_origin      complement(1103..1691)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1844..2146)
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRIVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(2305..2536)
                     /label=attP2
                     /note="recombination site for the Gateway(R) BP reaction 
                     (pDONR(TM)221 version)"
     promoter        complement(2554..2572)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2584..2600)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(2674..4092)
                     /codon_start=1
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     promoter        complement(4093..4538)
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             complement(4676..5332)
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        complement(5385..5415)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     rep_origin      5442..5795
                     /label=oriR6Kgamma
                     /note="oriR6Kgamma"
     oriT            5937..6046
                     /label=oriT
                     /note="incP origin of transfer"
ORIGIN
        1 ataacttcgt atagcataca ttatacgaag ttatgtcgac tctagcatgg atctcgggga
       61 cgtctaacta ctaagcgaga gtagggaact gccaggcatc aaataaaacg aaaggctcag
      121 tcggaagact gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg
      181 acaaatccgc cgggagcgga tttgaacgtt gtgaagcaac ggcccggagg gtggcgggca
      241 ggacgcccgc cataaactgc caggcatcaa actaagcaga aggccatcct gacggatggc
      301 ctttttgcgt ttctacaaac tcttcctgtt agttagttac ttaagcgcca gggttttccc
      361 agtcacgacg ttgtaaaacg acggccagtg catacgattt aggtgacact atagaagggc
      421 cccaaataat gattttattt tgactgatag tgacctgttc gttgcaacaa attgatgagc
      481 aatgcttttt tataatgcca actttgtaca aaaaagctga acgagaaacg taaaatgata
      541 taaatatcaa tatattaaat tagattttgc ataaaaaaca gactacataa tactgtaaaa
      601 cacaacatat ccagtcacta tgaatcaact acttagatgg tattagtgac ctgtagtcga
      661 ggcgcgccga ctttaggtcg aattccgcta gccttaagta actaactaac aggaagagtt
      721 tgtagaaacg caaaaaggcc atccgtcagg atggccttct gcttagtttg atgcctggca
      781 gtttatggcg ggcgtcctgc ccgccaccct ccgggccgtt gcttcacaac gttcaaatcc
      841 gctcccggcg gatttgtcct actcaggaga gcgttcaccg acaaacaaca gataaaacga
      901 aaggcccagt cttccgactg agcctttcgt tttatttgat gcctggcagt tccctactct
      961 cgcttagtag ttagacgtcc ccgagatcca tgctagcggt aatacggtta tccacagaat
     1021 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     1081 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     1141 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     1201 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     1261 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     1321 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     1381 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     1441 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     1501 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     1561 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     1621 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     1681 aaggatctca agaagatcct ttgatctttt ctaccgggtc tgacgctcag tggaacgggg
     1741 cgggcggcgt tgtgacaatt taccgaacaa ctccgcggcc gggaagccga tctcggcttg
     1801 aacgaattgt taggtggcgg tacttgggtc gatatcagca ctatattccc cagaacatca
     1861 ggttaatggc gtttttgatg tcattttcgc ggtggctgag atcagccact tcttccccga
     1921 taacggagac cggcacactg gccatatcgg tggtcatcat gcgccagctt tcatccccga
     1981 tatgcaccac cgggtaaagt tcacgggaga ctttatctga cagcagacgt gcactggcca
     2041 gggggatcac tatccgtcgc ccgggcgtgt caataatatc actctgtaca tccacaaaca
     2101 gacgataacg gctctctctt ttataggtgt aaaccttaaa ctgcattcta gaagcacacc
     2161 tctttttgac atacttcggg tatacatatc agtatatatt cttataccgc aaaaatcagc
     2221 gcgcaaatac gcatactgtt atctggcttt tagtaagctc atgagggaag cttggaccta
     2281 aagtcgttaa ttaacgggct cgactacagg tcactaatac catctaagta gttgattcat
     2341 agtgactgga tatgttgtgt tttacagtat tatgtagtct gttttttatg caaaatctaa
     2401 tttaatatat tgatatttat atcattttac gtttctcgtt cagctttctt gtacaaagtt
     2461 ggcattataa gaaagcattg cttatcaatt tgttgcaacg aacaggtcac tatcagtcaa
     2521 aataaaatca ttatttgcca tccagctgca ggccctatag tgagtcgtat taccgtaatc
     2581 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc tcatggcggc cgccaatgcc
     2641 aataggatat cggcattttc ttttgcgttt ttatttgtta actgttaatt gtccttgttc
     2701 aaggatgctg tctttgacaa cagatgtttt cttgcctttg atgttcagca ggaagctagg
     2761 cgcaaacgtt gattgtttgt ctgcgtagaa tcctctgttt gtcatatagc ttgtaatcac
     2821 gacattgttt cctttcgctt gaggtacagc gaagtgtgag taagtaaagg ttacatcgtt
     2881 aggatcaaga tccattttta acacaaggcc agttttgttc agcggcttgt atgggccagt
     2941 taaagaatta gaaacataac caagcatgta aatatcgtta gacgtaatgc cgtcaatcgt
     3001 catttttgat ccgcgcgagt cagtgaacaa gtaccatttg ccgttcattt taaagacgtt
     3061 cgcgcgttca atttcatctg ttactgtgtt agatgcaatc agcggtttca tcactttttt
     3121 cagtgtgtaa tcatcgttta gctcaatcat accgagagcg ccgtttgcta actcagccgt
     3181 gcgtttttta tcgctttgca gaagtttttg actttcttga cggaagaatg atgtgctttt
     3241 gccatagtat gctttgttaa ataaagattc ttcgccttgg tagccatctt cagttccagt
     3301 gtttgcttca aatactaagt atttgtggcc tttatcttct acgtagtgag gatctctcag
     3361 cgtatggttg tcgcctgagc tgtagttgcc ttcatcgatg aactgctgta cattttgata
     3421 cgtttttccg tcaccgtcaa agattgattt ataatcctct acaccgttga tgttcaaaga
     3481 gctgtctgat gctgatacgt taacttgtgc agttgtcagt gtttgtttgc cgtaatgttt
     3541 accggagaaa tcagtgtaga ataaacggat ttttccgtca gatgtaaatg tggctgaacc
     3601 tgaccattct tgtgtttggt cttttaggat agaatcattt gcatcgaatt tgtcgctgtc
     3661 tttaaagacg cggccagcgt ttttccagct gtcaatagaa gtttcgccga ctttttgata
     3721 gaacatgtaa atcgatgtgt catccgcatt tttaggatct ccggctaatg caaagacgat
     3781 gtggtagccg tgatagtttg cgacagtgcc gtcagcgttt tgtaatggcc agctgtccca
     3841 aacgtccagg ccttttgcag aagagatatt tttaattgtg gacgaatcga actcaggaac
     3901 ttgatatttt tcattttttt gctgttcagg gatttgcagc atatcatggc gtgtaatatg
     3961 ggaaatgccg tatgtttcct tatatggctt ttggttcgtt tctttcgcaa acgcttgagt
     4021 tgcgcctcct gccagcagtg cggtagtaaa ggttaatact gttgcttgtt ttgcaaactt
     4081 tttgatgttc atcgttcatg tctccttttt tatgtactgt gttagcggtc tgcttcttcc
     4141 agccctcctg tttgaagatg gcaagttagt tacgcacaat aaaaaaagac ctaaaatatg
     4201 taaggggtga cgccaaagta tacactttgc cctttacaca ttttaggtct tgcctgcttt
     4261 atcagtaaca aacccgcgcg atttactttt cgacctcatt ctattagact ctcgtttgga
     4321 ttgcaactgg tctattttcc tcttttgttt gatagaaaat cataaaagga tttgcagact
     4381 acgggcctaa agaactaaaa aatctatctg tttcttttca ttctctgtat tttttatagt
     4441 ttctgttgca tgggcataaa gttgcctttt taatcacaat tcagaaaata tcataatatc
     4501 tcatttcact aaataatagt gaacggcagg tatatgtggt cgagcctgat agtttggctg
     4561 tgagcaatta tgtgcttagt gcatctaatc gcttgagtta acgccggcga agcggcgtcg
     4621 gcttgaacga atttctagct agaggatcgc accaataact gccttaaaaa aattacgccc
     4681 cgccctgcca ctcatcgcag tactgttgta attcattaag cattctgccg acatggaagc
     4741 catcacaaac ggcatgatga acctgaatcg ccagcggcat cagcaccttg tcgccttgcg
     4801 tataatattt gcccattgtg aaaacggggg cgaagaagtt gtccatattg gccacgttta
     4861 aatcaaaact ggtgaaactc acccagggat tggctgagac gaaaaacata ttctcaataa
     4921 accctttagg gaaataggcc aggttttcac cgtaacacgc cacatcttgc gaatatatgt
     4981 gtagaaactg ccggaaatcg tcgtggtatt cactccagag cgatgaaaac gtttcagttt
     5041 gctcatggaa aacggtgtaa caagggtgaa cactatccca tatcaccagc tcaccgtctt
     5101 tcattgccat acggaactcc ggatgagcat tcatcaggcg ggcaagaatg tgaataaagg
     5161 ccggataaaa cttgtgctta tttttcttta cggtctttaa aaaggccgta atatccagct
     5221 gaacggtctg gttataggta cattgagcaa ctgactgaaa tgcctcaaaa tgttctttac
     5281 gatgccattg ggatatatca acggtggtat atccagtgat ttttttctcc attttagctt
     5341 ccttagctcc tgaaaatctc gacgatccta actcaaaatc cacacattat acgagccgga
     5401 agcataaagt gtaaagcctg ggggtgccta atgcgtctag ctgcgttaca tccctggctt
     5461 gttgtccaca accgttaaac cttaacagct ttaaaagcct tatatattct tttttttctt
     5521 ataaaactta aaaccttaga ggctatttaa gttgctgatt tatattaatt ttattgttca
     5581 aacatgagag cttagtacgt gaaacatgag agcttagtac gttagccatg agagcttagt
     5641 acgttagcca tgagggttta gttcgttaaa catgagagct tagtacgtta aacatgagag
     5701 cttagtacgt gaaacatgag agcttagtac gtactatcaa caggttgaac tgctgatctt
     5761 cagatcctct acgccggacg catcgtggcc ggatcaattc cgttttccgc tgcataaccc
     5821 tgcttcgggg tcattatagc gattttttcg gtatatccat cctttttcgc acgatataca
     5881 ggattttgcc aaagggttcg tgtagacttt ccttggtgta tccaacggcg tcagccgggc
     5941 aggataggtg aagtaggccc acccgcgagc gggtgttcct tcttcactgt cccttattcg
     6001 cacctggcgg tgctcaacgg gaatcctgct ctgcgaggct ggccggctac cgccggcgta
     6061 acagatgagg gcaagcggat ggctgatgaa accaagccaa ccaggaaggg cagcccacct
     6121 atcatcatgg
//
