LOCUS       40924_4100        8306 bp DNA     circular SYN 13-MAY-2021
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8306)
  TITLE     S cerevisiae Advanced Gateway Destination Vectors
REFERENCE   2  (bases 1 to 8306)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 8306)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8306
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward
                     primer"
     rep_origin      69..1411
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     promoter        1438..1542
                     /label=AmpR promoter
     CDS             1543..2400
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      2574..3162
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3316..3333
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3450..3471
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        3486..3516
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3524..3540
                     /label=lac operator
                     /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     3548..3564
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3585..3603
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3628..4271
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     4277..4293
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4301..4425
                     /label=attR1
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        4450..4480
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             4534..5190
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             5535..5837
                     /codon_start=1
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(5881..6005)
                     /label=attR2
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(6038..6054)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      6057..6304
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     promoter        complement(6323..6341)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6351..6367)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      6508..6963
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     CDS             complement(7097..7897)
                     /codon_start=1
                     /label=URA3
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(7898..8113)
                     /label=URA3 promoter
     primer_bind     complement(8137..8156)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable
                     marker"
     primer_bind     8256..8278
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagtatga tccaatatca aaggaaatga tagcattgaa ggatgagact aatccaattg
      121 aggagtggca gcatatagaa cagctaaagg gtagtgctga aggaagcata cgataccccg
      181 catggaatgg gataatatca caggaggtac tagactacct ttcatcctac ataaatagac
      241 gcatataagt acgcatttaa gcataaacac gcactatgcc gttcttctca tgtatatata
      301 tatacaggca acacgcagat ataggtgcga cgtgaacagt gagctgtatg tgcgcagctc
      361 gcgttgcatt ttcggaagcg ctcgttttcg gaaacgcttt gaagttccta ttccgaagtt
      421 cctattctct agaaagtata ggaacttcag agcgcttttg aaaaccaaaa gcgctctgaa
      481 gacgcacttt caaaaaacca aaaacgcacc ggactgtaac gagctactaa aatattgcga
      541 ataccgcttc cacaaacatt gctcaaaagt atctctttgc tatatatctc tgtgctatat
      601 ccctatataa cctacccatc cacctttcgc tccttgaact tgcatctaaa ctcgacctct
      661 acatttttta tgtttatctc tagtattact ctttagacaa aaaaattgta gtaagaacta
      721 ttcatagagt gaatcgaaaa caatacgaaa atgtaaacat ttcctatacg tagtatatag
      781 agacaaaata gaagaaaccg ttcataattt tctgaccaat gaagaatcat caacgctatc
      841 actttctgtt cacaaagtat gcgcaatcca catcggtata gaatataatc ggggatgcct
      901 ttatcttgaa aaaatgcacc cgcagcttcg ctagtaatca gtaaacgcgg gaagtggagt
      961 caggcttttt ttatggaaga gaaaatagac accaaagtag ccttcttcta accttaacgg
     1021 acctacagtg caaaaagtta tcaagagact gcattataga gcgcacaaag gagaaaaaaa
     1081 gtaatctaag atgctttgtt agaaaaatag cgctctcggg atgcattttt gtagaacaaa
     1141 aaagaagtat agattctttg ttggtaaaat agcgctctcg cgttgcattt ctgttctgta
     1201 aaaatgcagc tcagattctt tgtttgaaaa attagcgctc tcgcgttgca tttttgtttt
     1261 acaaaaatga agcacagatt cttcgttggt aaaatagcgc tttcgcgttg catttctgtt
     1321 ctgtaaaaat gcagctcaga ttctttgttt gaaaaattag cgctctcgcg ttgcattttt
     1381 gttctacaaa atgaagcaca gatgcttcgt tcaggtggca cttttcgggg aaatgtgcgc
     1441 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     1501 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     1561 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     1621 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     1681 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     1741 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     1801 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     1861 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     1921 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     1981 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     2041 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     2101 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     2161 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     2221 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     2281 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     2341 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     2401 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     2461 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     2521 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     2581 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     2641 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     2701 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     2761 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     2821 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     2881 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     2941 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     3001 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     3061 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     3121 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     3181 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     3241 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     3301 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     3361 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     3421 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     3481 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     3541 tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa ccctcactaa
     3601 agggaacaaa agctggagct cagtttatca ttatcaatac tcgccatttc aaagaatacg
     3661 taaataatta atagtagtga ttttcctaac tttatttagt caaaaaatta gccttttaat
     3721 tctgctgtaa cccgtacatg cccaaaatag ggggcgggtt acacagaata tataacatcg
     3781 taggtgtctg ggtgaacagt ttattcctgg catccactaa atataatgga gcccgctttt
     3841 taagctggca tccagaaaaa aaaagaatcc cagcaccaaa atattgtttt cttcaccaac
     3901 catcagttca taggtccatt ctcttagcgc aactacagag aacaggggca caaacaggca
     3961 aaaaacgggc acaacctcaa tggagtgatg caacctgcct ggagtaaatg atgacacaag
     4021 gcaattgacc cacgcatgta tctatctcat tttcttacac cttctattac cttctgctct
     4081 ctctgatttg gaaaaagctg aaaaaaaagg ttgaaaccag ttccctgaaa ttattcccct
     4141 acttgactaa taagtatata aagacggtag gtattgattg taattctgta aatctatttc
     4201 ttaaacttct taaattctac ttttatagtt agtctttttt ttagttttaa aacaccagaa
     4261 cttagtttcg acggattcta gaactagtgg atcccccatc acaagtttgt acaaaaaagc
     4321 tgaacgagaa acgtaaaatg atataaatat caatatatta aattagattt tgcataaaaa
     4381 acagactaca taatactgta aaacacaaca tatccagtca ctatggcggc cgcattaggc
     4441 accccaggct ttacacttta tgcttccggc tcgtataatg tgtggatttt gagttaggat
     4501 ccgtcgagat tttcaggagc taaggaagct aaaatggaga aaaaaatcac tggatatacc
     4561 accgttgata tatcccaatg gcatcgtaaa gaacattttg aggcatttca gtcagttgct
     4621 caatgtacct ataaccagac cgttcagctg gatattacgg cctttttaaa gaccgtaaag
     4681 aaaaataagc acaagtttta tccggccttt attcacattc ttgcccgcct gatgaatgct
     4741 catccggaat tccgtatggc aatgaaagac ggtgagctgg tgatatggga tagtgttcac
     4801 ccttgttaca ccgttttcca tgagcaaact gaaacgtttt catcgctctg gagtgaatac
     4861 cacgacgatt tccggcagtt tctacacata tattcgcaag atgtggcgtg ttacggtgaa
     4921 aacctggcct atttccctaa agggtttatt gagaatatgt ttttcgtctc agccaatccc
     4981 tgggtgagtt tcaccagttt tgatttaaac gtggccaata tggacaactt cttcgccccc
     5041 gttttcacca tgggcaaata ttatacgcaa ggcgacaagg tgctgatgcc gctggcgatt
     5101 caggttcatc atgccgtttg tgatggcttc catgtcggca gaatgcttaa tgaattacaa
     5161 cagtactgcg atgagtggca gggcggggcg taaacgcgtg gatccggctt actaaaagcc
     5221 agataacagt atgcgtattt gcgcgctgat ttttgcggta taagaatata tactgatatg
     5281 tatacccgaa gtatgtcaaa aagaggtatg ctatgaagca gcgtattaca gtgacagttg
     5341 acagcgacag ctatcagttg ctcaaggcat atatgatgtc aatatctccg gtctggtaag
     5401 cacaaccatg cagaatgaag cccgtcgtct gcgtgccgaa cgctggaaag cggaaaatca
     5461 ggaagggatg gctgaggtcg cccggtttat tgaaatgaac ggctcttttg ctgacgagaa
     5521 caggggctgg tgaaatgcag tttaaggttt acacctataa aagagagagc cgttatcgtc
     5581 tgtttgtgga tgtacagagt gatattattg acacgcccgg gcgacggatg gtgatccccc
     5641 tggccagtgc acgtctgctg tcagataaag tctcccgtga actttacccg gtggtgcata
     5701 tcggggatga aagctggcgc atgatgacca ccgatatggc cagtgtgccg gtctccgtta
     5761 tcggggaaga agtggctgat ctcagccacc gcgaaaatga catcaaaaac gccattaacc
     5821 tgatgttctg gggaatataa atgtcaggct cccttataca cagccagtct gcaggtcgac
     5881 catagtgact ggatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc
     5941 taatttaata tattgatatt tatatcattt tacgtttctc gttcagcttt cttgtacaaa
     6001 gtggtgatgg gctgcaggaa ttcgatatca agcttatcga taccgtcgac ctcgagtcat
     6061 gtaattagtt atgtcacgct tacattcacg ccctcccccc acatccgctc taaccgaaaa
     6121 ggaaggagtt agacaacctg aagtctaggt ccctatttat ttttttatag ttatgttagt
     6181 attaagaacg ttatttatat ttcaaatttt tctttttttt ctgtacagac gcgtgtacgc
     6241 atgtaacatt atactgaaaa ccttgcttga gaaggttttg ggacgctcga aggctttaat
     6301 ttgcggccgg tacccaattc gccctatagt gagtcgtatt acgcgcgctc actggccgtc
     6361 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
     6421 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
     6481 cagttgcgca gcctgaatgg cgaatggacg cgccctgtag cggcgcatta agcgcggcgg
     6541 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     6601 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     6661 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     6721 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     6781 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     6841 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     6901 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     6961 tttcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatagggta
     7021 ataactgata taattaaatt gaagctctaa tttgtgagtt tagtatacat gcatttactt
     7081 ataatacagt tttttagttt tgctggccgc atcttctcaa atatgcttcc cagcctgctt
     7141 ttctgtaacg ttcaccctct accttagcat cccttccctt tgcaaatagt cctcttccaa
     7201 caataataat gtcagatcct gtagagacca catcatccac ggttctatac tgttgaccca
     7261 atgcgtctcc cttgtcatct aaacccacac cgggtgtcat aatcaaccaa tcgtaacctt
     7321 catctcttcc acccatgtct ctttgagcaa taaagccgat aacaaaatct ttgtcgctct
     7381 tcgcaatgtc aacagtaccc ttagtatatt ctccagtaga tagggagccc ttgcatgaca
     7441 attctgctaa catcaaaagg cctctaggtt cctttgttac ttcttctgcc gcctgcttca
     7501 aaccgctaac aatacctggg cccaccacac cgtgtgcatt cgtaatgtct gcccattctg
     7561 ctattctgta tacacccgca gagtactgca atttgactgt attaccaatg tcagcaaatt
     7621 ttctgtcttc gaagagtaaa aaattgtact tggcggataa tgcctttagc ggcttaactg
     7681 tgccctccat ggaaaaatca gtcaagatat ccacatgtgt ttttagtaaa caaattttgg
     7741 gacctaatgc ttcaactaac tccagtaatt ccttggtggt acgaacatcc aatgaagcac
     7801 acaagtttgt ttgcttttcg tgcatgatat taaatagctt ggcagcaaca ggactaggat
     7861 gagtagcagc acgttcctta tatgtagctt tcgacatgat ttatcttcgt ttcctgcagg
     7921 tttttgttct gtgcagttgg gttaagaata ctgggcaatt tcatgtttct tcaacactac
     7981 atatgcgtat atataccaat ctaagtctgt gctccttcct tcgttcttcc ttctgttcgg
     8041 agattaccga atcaaaaaaa tttcaaggaa accgaaatca aaaaaaagaa taaaaaaaaa
     8101 atgatgaatt gaaaaggtgg tatggtgcac tctcagtaca atctgctctg atgccgcata
     8161 gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     8221 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     8281 ttcaccgtca tcaccgaaac gcgcga
//
