LOCUS       40924_35449        6169 bp DNA     circular SYN 18-DEC-2018
DEFINITION  MISSA donor vector pPr-ccdB, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6169)
  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 6169)
  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 6169)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6169)
  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..6169
                     /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    complement(429..660)
                     /label=attP2
                     /note="recombination site for the Gateway(R) BP reaction 
                     (pDONR(TM)221 version)"
     terminator      complement(728..755)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(847..933)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     rep_origin      complement(1098..1686)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1839..2141)
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRIVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    2346..2577
                     /label=attP1
                     /note="recombination site for the Gateway(R) BP reaction"
     promoter        complement(2593..2611)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2623..2639)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(2713..4131)
                     /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(4132..4577)
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             complement(4715..5371)
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        complement(5424..5454)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     rep_origin      5481..5834
                     /label=oriR6Kgamma
                     /note="oriR6Kgamma"
     oriT            5976..6085
                     /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 ccctcgacta caggtcacta ataccatcta agtagttgat tcatagtgac tggatatgtt
      481 gtgttttaca gtattatgta gtctgttttt tatgcaaaat ctaatttaat atattgatat
      541 ttatatcatt ttacgtttct cgttcagctt tcttgtacaa agttggcatt ataagaaagc
      601 attgcttatc aatttgttgc aacgaacagg tcactatcag tcaaaataaa atcattattt
      661 gccatccaga ggtcgaattc cgctagcctt aagtaactaa ctaacaggaa gagtttgtag
      721 aaacgcaaaa aggccatccg tcaggatggc cttctgctta gtttgatgcc tggcagttta
      781 tggcgggcgt cctgcccgcc accctccggg ccgttgcttc acaacgttca aatccgctcc
      841 cggcggattt gtcctactca ggagagcgtt caccgacaaa caacagataa aacgaaaggc
      901 ccagtcttcc gactgagcct ttcgttttat ttgatgcctg gcagttccct actctcgctt
      961 agtagttaga cgtccccgag atccatgcta gcggtaatac ggttatccac agaatcaggg
     1021 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     1081 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     1141 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     1201 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     1261 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     1321 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     1381 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     1441 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     1501 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     1561 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     1621 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     1681 tctcaagaag atcctttgat cttttctacc gggtctgacg ctcagtggaa cggggcgggc
     1741 ggcgttgtga caatttaccg aacaactccg cggccgggaa gccgatctcg gcttgaacga
     1801 attgttaggt ggcggtactt gggtcgatat cagcactata ttccccagaa catcaggtta
     1861 atggcgtttt tgatgtcatt ttcgcggtgg ctgagatcag ccacttcttc cccgataacg
     1921 gagaccggca cactggccat atcggtggtc atcatgcgcc agctttcatc cccgatatgc
     1981 accaccgggt aaagttcacg ggagacttta tctgacagca gacgtgcact ggccaggggg
     2041 atcactatcc gtcgcccggg cgtgtcaata atatcactct gtacatccac aaacagacga
     2101 taacggctct ctcttttata ggtgtaaacc ttaaactgca ttctagaagc acacctcttt
     2161 ttgacatact tcgggtatac atatcagtat atattcttat accgcaaaaa tcagcgcgca
     2221 aatacgcata ctgttatctg gcttttagta agctcatgag ggaagcttgg acctctggca
     2281 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
     2341 tagccaaata atgattttat tttgactgat agtgacctgt tcgttgcaac aaattgatga
     2401 gcaatgcttt tttataatgc caactttgta caaaaaagct gaacgagaaa cgtaaaatga
     2461 tataaatatc aatatattaa attagatttt gcataaaaaa cagactacat aatactgtaa
     2521 aacacaacat atccagtcac tatgaatcaa ctacttagat ggtattagtg acctgtagtc
     2581 gaggctgcag gccctatagt gagtcgtatt accgtaatca tggtcatagc tgtttcctgt
     2641 gtgaaattgt tatccgctct catggcggcc gccaatgcca ataggatatc ggcattttct
     2701 tttgcgtttt tatttgttaa ctgttaattg tccttgttca aggatgctgt ctttgacaac
     2761 agatgttttc ttgcctttga tgttcagcag gaagctaggc gcaaacgttg attgtttgtc
     2821 tgcgtagaat cctctgtttg tcatatagct tgtaatcacg acattgtttc ctttcgcttg
     2881 aggtacagcg aagtgtgagt aagtaaaggt tacatcgtta ggatcaagat ccatttttaa
     2941 cacaaggcca gttttgttca gcggcttgta tgggccagtt aaagaattag aaacataacc
     3001 aagcatgtaa atatcgttag acgtaatgcc gtcaatcgtc atttttgatc cgcgcgagtc
     3061 agtgaacaag taccatttgc cgttcatttt aaagacgttc gcgcgttcaa tttcatctgt
     3121 tactgtgtta gatgcaatca gcggtttcat cacttttttc agtgtgtaat catcgtttag
     3181 ctcaatcata ccgagagcgc cgtttgctaa ctcagccgtg cgttttttat cgctttgcag
     3241 aagtttttga ctttcttgac ggaagaatga tgtgcttttg ccatagtatg ctttgttaaa
     3301 taaagattct tcgccttggt agccatcttc agttccagtg tttgcttcaa atactaagta
     3361 tttgtggcct ttatcttcta cgtagtgagg atctctcagc gtatggttgt cgcctgagct
     3421 gtagttgcct tcatcgatga actgctgtac attttgatac gtttttccgt caccgtcaaa
     3481 gattgattta taatcctcta caccgttgat gttcaaagag ctgtctgatg ctgatacgtt
     3541 aacttgtgca gttgtcagtg tttgtttgcc gtaatgttta ccggagaaat cagtgtagaa
     3601 taaacggatt tttccgtcag atgtaaatgt ggctgaacct gaccattctt gtgtttggtc
     3661 ttttaggata gaatcatttg catcgaattt gtcgctgtct ttaaagacgc ggccagcgtt
     3721 tttccagctg tcaatagaag tttcgccgac tttttgatag aacatgtaaa tcgatgtgtc
     3781 atccgcattt ttaggatctc cggctaatgc aaagacgatg tggtagccgt gatagtttgc
     3841 gacagtgccg tcagcgtttt gtaatggcca gctgtcccaa acgtccaggc cttttgcaga
     3901 agagatattt ttaattgtgg acgaatcgaa ctcaggaact tgatattttt catttttttg
     3961 ctgttcaggg atttgcagca tatcatggcg tgtaatatgg gaaatgccgt atgtttcctt
     4021 atatggcttt tggttcgttt ctttcgcaaa cgcttgagtt gcgcctcctg ccagcagtgc
     4081 ggtagtaaag gttaatactg ttgcttgttt tgcaaacttt ttgatgttca tcgttcatgt
     4141 ctcctttttt atgtactgtg ttagcggtct gcttcttcca gccctcctgt ttgaagatgg
     4201 caagttagtt acgcacaata aaaaaagacc taaaatatgt aaggggtgac gccaaagtat
     4261 acactttgcc ctttacacat tttaggtctt gcctgcttta tcagtaacaa acccgcgcga
     4321 tttacttttc gacctcattc tattagactc tcgtttggat tgcaactggt ctattttcct
     4381 cttttgtttg atagaaaatc ataaaaggat ttgcagacta cgggcctaaa gaactaaaaa
     4441 atctatctgt ttcttttcat tctctgtatt ttttatagtt tctgttgcat gggcataaag
     4501 ttgccttttt aatcacaatt cagaaaatat cataatatct catttcacta aataatagtg
     4561 aacggcaggt atatgtggtc gagcctgata gtttggctgt gagcaattat gtgcttagtg
     4621 catctaatcg cttgagttaa cgccggcgaa gcggcgtcgg cttgaacgaa tttctagcta
     4681 gaggatcgca ccaataactg ccttaaaaaa attacgcccc gccctgccac tcatcgcagt
     4741 actgttgtaa ttcattaagc attctgccga catggaagcc atcacaaacg gcatgatgaa
     4801 cctgaatcgc cagcggcatc agcaccttgt cgccttgcgt ataatatttg cccattgtga
     4861 aaacgggggc gaagaagttg tccatattgg ccacgtttaa atcaaaactg gtgaaactca
     4921 cccagggatt ggctgagacg aaaaacatat tctcaataaa ccctttaggg aaataggcca
     4981 ggttttcacc gtaacacgcc acatcttgcg aatatatgtg tagaaactgc cggaaatcgt
     5041 cgtggtattc actccagagc gatgaaaacg tttcagtttg ctcatggaaa acggtgtaac
     5101 aagggtgaac actatcccat atcaccagct caccgtcttt cattgccata cggaactccg
     5161 gatgagcatt catcaggcgg gcaagaatgt gaataaaggc cggataaaac ttgtgcttat
     5221 ttttctttac ggtctttaaa aaggccgtaa tatccagctg aacggtctgg ttataggtac
     5281 attgagcaac tgactgaaat gcctcaaaat gttctttacg atgccattgg gatatatcaa
     5341 cggtggtata tccagtgatt tttttctcca ttttagcttc cttagctcct gaaaatctcg
     5401 acgatcctaa ctcaaaatcc acacattata cgagccggaa gcataaagtg taaagcctgg
     5461 gggtgcctaa tgcgtctagc tgcgttacat ccctggcttg ttgtccacaa ccgttaaacc
     5521 ttaacagctt taaaagcctt atatattctt ttttttctta taaaacttaa aaccttagag
     5581 gctatttaag ttgctgattt atattaattt tattgttcaa acatgagagc ttagtacgtg
     5641 aaacatgaga gcttagtacg ttagccatga gagcttagta cgttagccat gagggtttag
     5701 ttcgttaaac atgagagctt agtacgttaa acatgagagc ttagtacgtg aaacatgaga
     5761 gcttagtacg tactatcaac aggttgaact gctgatcttc agatcctcta cgccggacgc
     5821 atcgtggccg gatcaattcc gttttccgct gcataaccct gcttcggggt cattatagcg
     5881 attttttcgg tatatccatc ctttttcgca cgatatacag gattttgcca aagggttcgt
     5941 gtagactttc cttggtgtat ccaacggcgt cagccgggca ggataggtga agtaggccca
     6001 cccgcgagcg ggtgttcctt cttcactgtc ccttattcgc acctggcggt gctcaacggg
     6061 aatcctgctc tgcgaggctg gccggctacc gccggcgtaa cagatgaggg caagcggatg
     6121 gctgatgaaa ccaagccaac caggaagggc agcccaccta tcatcatgg
//
