LOCUS       pCMV5D_HA_PPM1M_PPM1H_F 6400 bp ds-DNA     circular     15-JAN-2024
DEFINITION  .
KEYWORDS    "creator:MRCPPU" "marker:HygR, AmpR"
FEATURES             Location/Qualifiers
     enhancer        235..614
                     /label="CMV enhancer"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label="CMV promoter"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: human cytomegalovirus (CMV) immediate early promoter"
     CDS             863..889
                     /label="HA"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
                     /note="codon_start: 1 product: HA (human influenza hemagglutinin) epitope tag transl_table: 1 translation: YPYDVPDYA"
     CDS             863..889
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="YPYDVPDYA"
     primer          875..912
                     /label="PPM1M_CSII_mApple_GA_F"
                     /note="sequence: GAACACGCTACCGGTCTCGAGATGTCCGCCGGCTGGTT"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     CDS             890..895
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="GS"
     CDS             896..2275
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MSAGWFRRRFLPGEPLPAPRPPGPHASPVPYRRPRFLRGSSSSPGAADASRRPDSRPVRSPARGRTLPWNAGYAEIINAEKSEFNEDQAACGKLCIRRCEFGAEEEWLTLCPEEFLTGHYWALFDGHGGPAAAILAANTLHSCLRRQLEAVVEGLVATQPPMHLNGRCICPSDPQFVEEKGIRAEDLVIGALESAFQECDEVIGRELEASGQMGGCTALVAVSLQGKLYMANAGDSRAILVRRDEIRPLSSEFTPETERQRLQYLAFMQPHLLGNEFTHLEFPRRVQRKELGKKMLYRDFNMTGWAYKTIEDEDLKFPLIYGEGKKARVMATIGVTRGLGDHDLKVHDSNIYIKPFLSSAPQVTVLDVDQLELQEDDVVVMATDGLWDVLSNEQVAWLVRSFLPGNQEDPHRFSKLAQMLIHSTQGKEDSLTEEGQVSYDDVSVFVIPLHSQGQESSDH*"
     misc_feature    896..2275
                     /label="PPM1M"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
     primer          complement(1260..1284)
                     /label="PPM1M H127D GA R"
                     /note="sequence: GGACCGCCGTCCCCATCGAACAGTG"
                     /ApEinfo_revcolor="#85dae9"
                     /ApEinfo_fwdcolor="#85dae9"
     primer          1264..1287
                     /label="PPM1M H127D GA F"
                     /note="sequence: GTTCGATGGGGACGGCGGTCCTGC"
                     /ApEinfo_revcolor="#c7b0e3"
                     /ApEinfo_fwdcolor="#c7b0e3"
     misc_feature    1274..1276
                     /label="H127D"
                     /ApEinfo_revcolor="#75c6a9"
                     /ApEinfo_fwdcolor="#75c6a9"
     primer          complement(1583..1607)
                     /label="PPM1M D235A GA R"
                     /note="sequence: CCCTGCTAGCCCCAGCATTGGCCAT"
                     /ApEinfo_revcolor="#85dae9"
                     /ApEinfo_fwdcolor="#85dae9"
     primer          1588..1612
                     /label="PPM1M D235A GA F"
                     /note="sequence: CAATGCTGGGGCTAGCAGGGCCATC"
                     /ApEinfo_revcolor="#75c6a9"
                     /ApEinfo_fwdcolor="#75c6a9"
     misc_feature    1598..1600
                     /label="D235A"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
     misc_feature    1646..1978
                     /label="PPM1H FLAP"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
     primer          complement(2252..2291)
                     /label="PPM1M_CSII_mApple_GA_R"
                     /note="sequence: TGCTCACCATCAGAATTCCGTGGTCACTGCTCTCTTGGCC"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
     CDS             2276..2327
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="AAAYPYDVPDYA*VESR"
     CDS             2285..2311
                     /label="HA"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
                     /note="codon_start: 1 product: HA (human influenza hemagglutinin) epitope tag transl_table: 1 translation: YPYDVPDYA"
     polyA_signal    2358..2582
                     /label="bGH poly(A) signal"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
                     /note="note: bovine growth hormone polyadenylation signal"
     protein_bind    2866..2913
                     /label="FRT"
                     /ApEinfo_revcolor="#31849b"
                     /ApEinfo_fwdcolor="#31849b"
                     /note="bound_moiety: FLP recombinase from the Saccharomyces cerevisiae 2μ plasmid note: FLP-mediated recombination occurs in the 8-bp core sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2921..3943
                     /label="HygR"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
                     /note="codon_start: 1 gene: aph(4)-Ia note: confers resistance to hygromycin product: aminoglycoside phosphotransferase from E. coli transl_table: 1 translation: KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE*"
     polyA_signal    4073..4194
                     /label="SV40 poly(A) signal"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
                     /note="note: SV40 polyadenylation signal"
     primer_bind     complement(4243..4259)
                     /label="M13 rev"
                     /ApEinfo_revcolor="#a020f0"
                     /ApEinfo_fwdcolor="#a020f0"
                     /note="note: common sequencing primer, one of multiple similar variants"
     protein_bind    4267..4283
                     /label="lac operator"
                     /ApEinfo_revcolor="#31849b"
                     /ApEinfo_fwdcolor="#31849b"
                     /note="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-β-D-thiogalactopyranoside (IPTG)."
     promoter        complement(4291..4297)
                     /label="lac promoter"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: promoter for the E. coli lac operon"
     promoter        complement(4298..4315)
                     /label="lac promoter"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: promoter for the E. coli lac operon"
     promoter        complement(4316..4321)
                     /label="lac promoter"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: promoter for the E. coli lac operon"
     protein_bind    4336..4357
                     /label="CAP binding site"
                     /ApEinfo_revcolor="#31849b"
                     /ApEinfo_fwdcolor="#31849b"
                     /note="bound_moiety: E. coli catabolite activator protein note: CAP binding activates transcription in the presence of cAMP."
     rep_origin      complement(4645..5233)
                     /label="ori"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
                     /note="direction: LEFT note: high-copy-number ColE1/pMB1/pBR322/pUC origin of replication"
     CDS             complement(5404..6195)
                     /label="AmpR"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
                     /note="codon_start: 1 gene: bla note: confers resistance to ampicillin, carbenicillin, and related antibiotics product: β-lactamase transl_table: 1 translation: MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW*"
     CDS             complement(6196..6264)
                     /label="AmpR"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
                     /note="codon_start: 1 gene: bla note: confers resistance to ampicillin, carbenicillin, and related antibiotics product: β-lactamase transl_table: 1 translation: MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW*"
     promoter        complement(6265..6369)
                     /label="AmpR promoter"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="gene: bla"
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtccgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta cgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacaccaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta ataaccccgc cccgttgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagaatt
      841 ggccaccgga tctgccgcca tgtacccata cgatgtgcca gattacgccG GATCCATGTC
      901 CGCCGGCTGG TTCCGGCGCC GCTTCCTGCC TGGGGAGCCG CTCCCCGCGC CGCGTCCACC
      961 TGGGCCGCAT GCCAGCCCCG TGCCCTACCG ACGGCCCCGC TTCCTTCGCG GCTCCAGCTC
     1021 CAGCCCCGGG GCGGCCGACG CCTCGCGCCG CCCAGACTCC CGGCCCGTGC GCAGCCCCGC
     1081 ACGAGGACGC ACGCTACCCT GGAATGCAGG CTACGCCGAG ATTATCAATG CAGAGAAATC
     1141 TGAATTCAAT GAGGATCAAG CCGCCTGTGG GAAGCTGTGC ATCCGGAGAT GTGAGTTTGG
     1201 GGCTGAAGAA GAGTGGCTGA CCCTGTGCCC AGAGGAGTTC CTGACAGGCC ATTACTGGGC
     1261 ACTGTTCGAT GGGCACGGCG GTCCTGCAGC AGCCATCTTG GCTGCCAACA CCCTGCACTC
     1321 CTGCTTGCGC CGGCAGCTGG AGGCCGTGGT GGAAGGCTTG GTGGCCACTC AGCCCCCCAT
     1381 GCACCTCAAT GGCCGCTGCA TCTGCCCCAG TGACCCTCAG TTTGTGGAGG AAAAGGGCAT
     1441 CAGGGCAGAA GACTTGGTGA TCGGGGCATT GGAGAGTGCC TTTCAGGAAT GTGATGAGGT
     1501 GATCGGGCGG GAGCTGGAGG CCTCAGGCCA GATGGGCGGC TGCACAGCCC TGGTGGCTGT
     1561 GTCCCTGCAG GGAAAGCTGT ACATGGCCAA TGCTGGGGAT AGCAGGGCCA TCTTGGTGCG
     1621 GAGAGATGAG ATACGGCCAC TGAGCTCAGA ATTTACCCCC GAGACGGAGC GCCAGCGACT
     1681 TCAGTACCTG GCATTCATGC AGCCTCACTT GCTGGGAAAT GAGTTCACAC ATTTGGAGTT
     1741 TCCAAGGAGA GTACAGAGAA AGGAGCTTGG AAAGAAGATG CTCTACAGGG ACTTTAATAT
     1801 GACAGGCTGG GCATACAAAA CCATTGAGGA TGAGGACTTG AAGTTCCCCC TTATATATGG
     1861 AGAAGGCAAG AAGGCCCGGG TAATGGCAAC TATTGGAGTG ACCAGGGGAC TTGGGGACCA
     1921 TGACCTGAAG GTGCATGACT CCAACATCTA CATTAAACCA TTCCTGTCTT CAGCTCCACA
     1981 GGTGACTGTG CTGGATGTGG ACCAGCTGGA GCTACAGGAG GATGATGTGG TTGTCATGGC
     2041 AACTGATGGA CTCTGGGATG TACTGTCCAA CGAGCAGGTG GCATGGCTGG TGCGGAGCTT
     2101 CCTCCCTGGG AACCAAGAGG ACCCACACAG GTTCTCAAAG CTGGCCCAGA TGCTGATACA
     2161 CAGCACACAG GGAAAGGAAG ACAGTCTCAC AGAGGAAGGG CAGGTGTCCT ACGATGACGT
     2221 CTCTGTGTTC GTGATTCCCT TGCACAGTCA GGGCCAAGAG AGCAGTGACC ACTGAGCGGC
     2281 CGCttaccca tacgatgtgc cagattacgc ctaagtcgag tctagagggc ccgtttaaac
     2341 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     2401 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     2461 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     2521 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     2581 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     2641 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     2701 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     2761 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     2821 cctcgacccc aaaaaacttg attagggtga tggttcacgt acctagaagt tcctattccg
     2881 aagttcctat tctctagaaa gtataggaac ttccttggcc aaaaagcctg aactcaccgc
     2941 gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct
     3001 ctcggagggc gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct
     3061 gcgggtaaat agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc
     3121 atcggccgcg ctcccgattc cggaagtgct tgacattggg gagttcagcg agagcctgac
     3181 ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact
     3241 gcccgctgtt ctgcagccgg tcgcggaggc catggatgcg atcgctgcgg ccgatcttag
     3301 ccagacgagc gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg
     3361 tgatttcata tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga
     3421 caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg
     3481 ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa
     3541 tggccgcata acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga
     3601 ggtcgccaac atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta
     3661 cttcgagcgg aggcatccgg agcttgcagg atcgccgcgg ctccgggcgt atatgctccg
     3721 cattggtctt gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg
     3781 ggcgcagggt cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca
     3841 aatcgcccgc agaagcgcgg ccgtctggac cgatggctgt gtagaagtac tcgccgatag
     3901 tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt
     3961 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     4021 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     4081 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4141 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     4201 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     4261 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     4321 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4381 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4441 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4501 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4561 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4621 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4681 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4741 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4801 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4861 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4921 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4981 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     5041 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     5101 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5161 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     5221 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5281 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5341 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5401 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5461 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5521 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5581 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5641 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5701 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5761 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5821 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5881 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5941 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     6001 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     6061 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6121 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6181 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6241 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6301 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6361 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
//