LOCUS       pml248_pGP8GF-ccdB      7247 bp ds-DNA     circular     16-JUN-2022
DEFINITION  .
FEATURES             Location/Qualifiers
     misc_feature    complement(211..233)
                     /label="1483"
     primer          complement(211..233)
                     /label="1483"
                     /note="sequence: gggggatcatgtaactcgccttg"
     misc_feature    214..236
                     /label="1482"
     primer          214..236
                     /label="1482"
                     /note="sequence: ggcgagttacatgatcccccatg"
     promoter        complement(692..790)
                     /label="bla promoter"
     promoter        complement(733..761)
                     /label="AmpR_promoter"
     primer_bind     complement(910..932)
                     /label="3GEX"
     misc_feature    complement(910..932)
                     /label="pGEX_3_primer"
     misc_feature    1066..1071
                     /label="unused HIS3 5' UTR homology"
     misc_feature    1072..1079
                     /label="PmeI site"
     misc_feature    1080..1391
                     /label="HIS3 5' UTR"
     misc_feature    1392..2054
                     /label="HIS3"
     misc_feature    2055..2236
                     /label="ADH1 Terminator"
     primer_bind     complement(2207..2231)
                     /label="pGILDA-R"
     primer_bind     2237..2275
                     /label="oNL1872 GPDf"
     misc_feature    2237..2891
                     /label="Actual GPD Promoter"
     misc_feature    2780..2803
                     /label="GPDPro_F_primer (2002)"
     misc_feature    complement(2888..2923)
                     /label="3679"
     misc_feature    2892..2908
                     /label="pBluescriptSK_primer"
     misc_feature    complement(2895..2923)
                     /label="30"
     misc_feature    2916..2919
                     /label="attR1"
     misc_feature    2923..2930
                     /label="p2"
     misc_feature    2924..4337
                     /label="ccdB frag"
     promoter        2940..2963
                     /label="lac_promoter"
     CDS             3024..3607
                     /label="ORF frame 2"
     gene            3024..3607
                     /label="CAT/CamR"
     CDS             3609..3680
                     /label="CmR"
     CDS             3609..3683
                     /label="ORF frame 2"
     gene            3609..3683
                     /label="CAT/CamR"
     misc_feature    4025..4242
                     /label="ccdB"
     primer_bind     complement(4221..4256)
                     /label="j5_0033_(ccdB1)_(CtoG)_reverse"
     primer_bind     4231..4266
                     /label="j5_0034_(ccdB2)_forward"
     misc_feature    4243..4243
                     /label="synth"
     misc_feature    4244..4330
                     /label="ccdB"
     primer          complement(4309..4349)
                     /label="3903"
                     /note="sequence: TCCACCAGATCCGGAGACCTTATATTCCCCAGAACATCAGG"
     primer          4320..4359
                     /label="3904"
                     /note="sequence: GGGGAATATAAGGTCTCCGGATCTGGTGGAGATTACAAGG"
     misc_feature    4338..4359
                     /label="3596"
     primer          4338..4365
                     /label="3821"
                     /note="sequence: ggatctggtggagattacaaggatcatg"
     CDS             4338..4436
                     /label="Translation 4338-4436"
     misc_feature    4338..4436
                     /label="flagtag frag"
     protein_bind    4350..4415
                     /label="3xFLAG"
     misc_binding    4416..4433
                     /label="6xHis"
     primer          4424..4466
                     /label="3902"
                     /note="sequence: TCACCACCATTGATTCTATGTCAGGCTCCCTTATACACAGCCA"
     misc_feature    4437..2923
                     /label="pGP8G2 frag"
     misc_feature    4481..4498
                     /label="attR2"
     misc_feature    complement(4481..4498)
                     /label="attR1"
     misc_feature    4499..4581
                     /label="attR2"
     misc_feature    complement(4499..4588)
                     /label="attR1"
     primer_bind     complement(4664..4693)
                     /label="oNL1913 TCYC1 front Rev"
     terminator      4664..4903
                     /label="CYC1_terminator"
     misc_feature    complement(4673..4691)
                     /label="pYESTrp_rev_primer"
     misc_feature    complement(4673..4691)
                     /label="CYC1_primer"
     primer_bind     complement(4880..4903)
                     /label="oNL1475 TCYC1r"
     misc_feature    4904..5904
                     /label="HIS3 3'UTR + DED1 homology"
     misc_feature    5905..5912
                     /label="PmeI site"
     misc_feature    complement(5913..5931)
                     /label="M13_reverse_primer"
     primer_bind     complement(5917..5933)
                     /label="M13R"
     misc_feature    complement(5930..5952)
                     /label="M13_pUC_rev_primer"
     misc_binding    complement(5939..5961)
                     /label="LacO"
     rep_origin      complement(6301..6983)
                     /label="ColE1 origin"
     rep_origin      complement(6304..6923)
                     /label="pBR322_origin"
     CDS             complement(7076..691)
                     /label="Translation 691-7076"
     gene            complement(7078..7221)
                     /label="Ampicillin"
     primer_bind     complement(7197..7235)
                     /label="j5_0066_(pGP8G1)_(CtoT)_reverse"
     primer_bind     7210..7240
                     /label="j5_0067_(pGP8G2-start)_forward"
     misc_feature    7222..7222
                     /label="p"
     CDS             complement(7223..691)
                     /label="Ampicillin"
ORIGIN
        1 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
       61 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
      121 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
      181 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
      241 gaaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
      301 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
      361 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
      421 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
      481 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
      541 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
      601 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
      661 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
      721 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
      781 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctta ttatcatgac
      841 attaacctat aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga
      901 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
      961 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg
     1021 gcttaactat gcggcatcag agcagattgt actgagagtg caccataatt cgtttaaacc
     1081 gttttaagag cttggtgagc gctaggagtc actgccaggt atcgtttgaa cacggcatta
     1141 gtcagggaag tcataacaca gtcctttccc gcaattttct ttttctatta ctcttggcct
     1201 cctctagtac actctatatt tttttatgcc tcggtaatga ttttcatttt tttttttcca
     1261 cctagcggat gactcttttt ttttcttagc gattggcatt atcacataat gaattataca
     1321 ttatataaag taatgtgatt tcttcgaaga atatactaaa aaatgagcag gcaagataaa
     1381 cgaaggcaaa gatgacagag cagaaagccc tagtaaagcg tattacaaat gaaaccaaga
     1441 ttcagattgc gatctcttta aagggtggtc ccctagcgat agagcactcg atcttcccag
     1501 aaaaagaggc agaagcagta gcagaacagg ccacacaatc gcaagtgatt aacgtccaca
     1561 caggtatagg gtttctggac catatgatac atgctctggc caagcattcc ggctggtcgc
     1621 taatcgttga gtgcattggt gacttacaca tagacgacca tcacaccact gaagactgcg
     1681 ggattgctct cggtcaagct tttaaagagg ccctaggggc cgtgcgtgga gtaaaaaggt
     1741 ttggatcagg atttgcgcct ttggatgagg cactttccag agcggtggta gatctttcga
     1801 acaggccgta cgcagttgtc gaacttggtt tgcaaaggga gaaagtagga gatctctctt
     1861 gcgagatgat cccgcatttt cttgaaagct ttgcagaggc tagcagaatt accctccacg
     1921 ttgattgtct gcgaggcaag aatgatcatc accgtagtga gagtgcgttc aaggctcttg
     1981 cggttgccat aagagaagcc acctcgccca atggtaccaa cgatgttccc tccaccaaag
     2041 gtgttcttat gtaggcgaat ttcttatgat ttatgatttt tattattaaa taagttataa
     2101 aaaaaataag tgtatacaaa ttttaaagtg actcttaggt tttaaaacga aaattcttat
     2161 tcttgagtaa ctctttcctg taggtcaggt tgctttctca ggtatagcat gaggtcgctc
     2221 ttattgacca cacctcagtt tatcattatc aatactcgcc atttcaaaga atacgtaaat
     2281 aattaatagt agtgattttc ctaactttat ttagtcaaaa aattagcctt ttaattctgc
     2341 tgtaacccgt acatgcccaa aatagggggc gggttacaca gaatatataa catcgtaggt
     2401 gtctgggtga acagtttatt cctggcatcc actaaatata atggagcccg ctttttaagc
     2461 tggcatccag aaaaaaaaag aatcccagca ccaaaatatt gttttcttca ccaaccatca
     2521 gttcataggt ccattctctt agcgcaacta cagagaacag gggcacaaac aggcaaaaaa
     2581 cgggcacaac ctcaatggag tgatgcaacc tgcctggagt aaatgatgac acaaggcaat
     2641 tgacccacgc atgtatctat ctcattttct tacaccttct attaccttct gctctctctg
     2701 atttggaaaa agctgaaaaa aaaggttgaa accagttccc tgaaattatt cccctacttg
     2761 actaataagt atataaagac ggtaggtatt gattgtaatt ctgtaaatct atttcttaaa
     2821 cttcttaaat tctactttta tagttagtct tttttttagt tttaaaacac cagaacttag
     2881 tttcgacgga ttctagaact agtggatccc ccatcacaaa atgggagacc accccaggct
     2941 ttacacttta tgcttccggc tcgtataatg tgtggatttt gagttaggat ccgtcgagat
     3001 tttcaggagc taaggaagct aaaatggaga aaaaaatcac tggatatacc accgttgata
     3061 tatcccaatg gcatcgtaaa gaacattttg aggcatttca gtcagttgct caatgtacct
     3121 ataaccagac cgttcagctg gatattacgg cctttttaaa gaccgtaaag aaaaataagc
     3181 acaagtttta tccggccttt attcacattc ttgcccgcct gatgaatgct catccggaat
     3241 tccgtatggc aatgaaagac ggtgagctgg tgatatggga tagtgttcac ccttgttaca
     3301 ccgttttcca tgagcaaact gaaacgtttt catcgctctg gagtgaatac cacgacgatt
     3361 tccggcagtt tctacacata tattcgcaag atgtggcgtg ttacggtgaa aacctggcct
     3421 atttccctaa agggtttatt gagaatatgt ttttcgtctc agccaatccc tgggtgagtt
     3481 tcaccagttt tgatttaaac gtggccaata tggacaactt cttcgccccc gttttcacca
     3541 tgggcaaata ttatacgcaa ggcgacaagg tgctgatgcc gctggcgatt caggttcatc
     3601 atgccgtttg tgatggcttc catgtcggca gaatgcttaa tgaattacaa cagtactgcg
     3661 atgagtggca gggcggggcg taaacgcgtg gatccggctt actaaaagcc agataacagt
     3721 atgcgtattt gcgcgctgat ttttgcggta taagaatata tactgatatg tatacccgaa
     3781 gtatgtcaaa aagaggtatg ctatgaagca gcgtattaca gtgacagttg acagcgacag
     3841 ctatcagttg ctcaaggcat atatgatgtc aatatctccg gtctggtaag cacaaccatg
     3901 cagaatgaag cccgtcgtct gcgtgccgaa cgctggaaag cggaaaatca ggaagggatg
     3961 gctgaggtcg cccggtttat tgaaatgaac ggctcttttg ctgacgagaa caggggctgg
     4021 tgaaatgcag tttaaggttt acacctataa aagagagagc cgttatcgtc tgtttgtgga
     4081 tgtacagagt gatattattg acacgcccgg gcgacggatg gtgatccccc tggccagtgc
     4141 acgtctgctg tcagataaag tctcccgtga actttacccg gtggtgcata tcggggatga
     4201 aagctggcgc atgatgacca ccgatatggc cagtgtgccg gtgtccgtta tcggggaaga
     4261 agtggctgat ctcagccacc gcgaaaatga catcaaaaac gccattaacc tgatgttctg
     4321 gggaatataa ggtctccgga tctggtggag attacaagga tcatgatggt gattacaagg
     4381 atcacgacat cgactacaag gatgacgatg acaagcacca tcatcaccac cattgattct
     4441 atgtcaggct cccttataca cagccagtct gcaggtcgac catagtgact ggatatgttg
     4501 tgttttacag tattatgtag tctgtttttt atgcaaaatc taatttaata tattgatatt
     4561 tatatcattt tacgtttctc gttcagcttt cttgtacaaa gtggtgatgg gctgcaggaa
     4621 ttcgatTtca agcttatcga taccgtcgac ctcgagtcat gtaattagtt atgtcacgct
     4681 tacattcacg ccctcccccc acatccgctc taaccgaaaa ggaaggagtt agacaacctg
     4741 aagtctaggt ccctatttat ttttttatag ttatgttagt attaagaacg ttatttatat
     4801 ttcaaatttt tctttttttt ctgtacagac gcgtgtacgc atgtaacatt atactgaaaa
     4861 ccttgcttga gaaggttttg ggacgctcga aggctttaat ttgtgacacc gattatttaa
     4921 agctgcagca tacgatatat atacatgtgt atatatgtat acctatgaat gtcagtaagt
     4981 atgtatacga acagtatgat actgaagatg acaaggtaat gcatcattct atacgtgtca
     5041 ttctgaacga ggcgcgcttt ccttttttct ttttgctttt tctttttttt tctcttgaac
     5101 tcgagaaaaa aaatataaaa gagatggagg aacgggaaaa agttagttgt ggtgataggt
     5161 ggcaagtggt attccgtaag aacaacaaga aaagcatttc atattatggc tgaactgagc
     5221 gaacaagtgc aaaatttaag catcaacgac aacaacgaga atggttatgt tcctcctcac
     5281 ttaagaggaa aaccaagaag tgccagaaat aacagtagca actacaataa caacaacggc
     5341 ggctacaacg gtggccgtgg cggtggcagc ttctttagca acaaccgtcg tggtggttac
     5401 ggcaacggtg gtttcttcgg tggaaacaac ggtggcagca gatctaacgg ccgttctggt
     5461 ggtagatgga tcgatggcaa acatgtccca gctccaagaa acgaaaaggc cgagatcgcc
     5521 atatttggtg tccccgagga tccaaatttc caatcttctg gtattaactt cgataactac
     5581 gatgatattc cagtggacgc ctctggtaag gatgttcctg aaccaatcac agaatttacc
     5641 tcacctccat tggacggatt gttattggaa aacatcaaat tggcccgttt caccaagcca
     5701 acacctgtgc aaaaatactc cgtccctatc gttgccaacg gcagagattt gatggcctgt
     5761 gcgcagaccg gttctggtaa gactggtggg tttttattcc cagtgttgtc cgaatcattt
     5821 aagactggac catctcctca accagagtct caaggctcct tttaccaaag aaaggcctac
     5881 ccaactgctg tcattatggc tccagtttaa accatggtca tagctgtttc ctgtgtgaaa
     5941 ttgttatccg ctcacaattc cacacaacat aggagccgga agcataaagt gtaaagcctg
     6001 gggtgcctaa tgagtgaggt aactcacatt aattgcgttg cgctcactgc ccgctttcca
     6061 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6121 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     6181 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     6241 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     6301 ggccgcgttg ctggcgtttt tccataggct cggcccccct gacgagcatc acaaaaatcg
     6361 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgttcccccc
     6421 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     6481 ctttctccct tcgggaagcg tggcgctttc tcaatgctca cgctgtaggt atctcagttc
     6541 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     6601 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     6661 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     6721 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     6781 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     6841 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     6901 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     6961 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     7021 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     7081 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     7141 tgcctgactg cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     7201 tgctgcaatg ataccgcgag atccacgctc accggctcca gatttat
//