LOCUS       sequence-429147-        7318 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human MPL(N358Q).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7318)
  TITLE     Masubuchi et al., 2024
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7318)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pcDNA3.1/MPL(N358Q)
FEATURES             Location/Qualifiers
     source          1..7318
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2909..2926)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2915..3139
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3185..3613
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3272..3291)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3482..3503
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3622..3642)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3627..3956
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3807..3942
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3869..3888
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4023..4817
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4077..4096)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4687..4706
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4991..5112
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5028..5047)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5082..5101
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5161..5177)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5161..5177)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5174..5196)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5185..5201
                     /label=lac operator
                     /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-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5209..5215,5216..5233,5234..5239))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5254..5275
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5392..5409)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5563..6151)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5643..5662)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6322..7113,7114..7182))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6945..6964
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7183..7287)
                     /gene="bla"
                     /label=AmpR promoter
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 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattcgcc
      961 accatgccct cctgggccct cttcatggtc acctcctgcc tcctcctggc ccctcaaaac
     1021 ctggcccaag tcagcagcca agatgtctcc ttgctggcat cagactcaga gcccctgaag
     1081 tgtttctccc gaacatttga ggacctcact tgcttctggg atgaggaaga ggcagcgccc
     1141 agtgggacat accagctgct gtatgcctac ccgcgggaga agccccgtgc ttgccccctg
     1201 agttcccaga gcatgcccca ctttggaacc cgatacgtgt gccagtttcc agaccaggag
     1261 gaagtgcgtc tcttctttcc gctgcacctc tgggtgaaga atgtgttcct aaaccagact
     1321 cggactcagc gagtcctctt tgtggacagt gtaggcctgc cggctccccc cagtatcatc
     1381 aaggccatgg gtgggagcca gccaggggaa cttcagatca gctgggagga gccagctcca
     1441 gaaatcagtg atttcctgag gtacgaactc cgctatggcc ccagagatcc caagaactcc
     1501 actggtccca cggtcataca gctgattgcc acagaaacct gctgccctgc tctgcagagg
     1561 cctcactcag cctctgctct ggaccagtct ccatgtgctc agcccacaat gccctggcaa
     1621 gatggaccaa agcagacctc cccaagtaga gaagcttcag ctctgacagc agagggtgga
     1681 agctgcctca tctcaggact ccagcctggc aactcctact ggctgcagct gcgcagcgaa
     1741 cctgatggga tctccctcgg tggctcctgg ggatcctggt ccctccctgt gactgtggac
     1801 ctgcctggag atgcagtggc acttggactg caatgcttta ccttggacct gaagaatgtt
     1861 acctgtcaat ggcagcaaca ggaccatgct agctcccaag gcttcttcta ccacagcagg
     1921 gcacggtgct gccccagaga caggtacccc atctgggaga actgcgaaga ggaagagaaa
     1981 acaaatccag gactacagac cccacagttc tctcgctgcc acttcaagtc acgacaggac
     2041 agcattattc acatccttgt ggaggtgacc acagccccgg gtactgttca cagctacctg
     2101 ggctcccctt tctggatcca ccaggctgtg cgcctcccca ccccaaactt gcactggagg
     2161 gagatctcca gtgggcatct ggaattggag tggcagcacc catcgtcctg ggcagcccaa
     2221 gagacctgtt atcaactccg atacacagga gaaggccatc aggactggaa ggtgctggag
     2281 ccgcctctcg gggcccgagg agggaccctg gagctgcgcc cgcgatctcg ctaccgttta
     2341 cagctgcgcg ccaggctcaa cggccccacc taccaaggtc cctggagctc gtggtcggac
     2401 ccaactaggg tggagaccgc caccgagacc gcctggatct ccttggtgac cgctctgcat
     2461 ctagtgctgg gcctcagcgc cgtcctgggc ctgctgctgc tgaggtggca gtttcctgca
     2521 cactacagga gactgaggca tgccctgtgg ccctcacttc cagacctgca ccgggtccta
     2581 ggccagtacc ttagggacac tgcagccctg agcccgccca aggccacagt ctcagatacc
     2641 tgtgaagaag tggaacccag cctccttgaa atcctcccca agtcctcaga gaggactcct
     2701 ttgcccctgt gttcctccca ggcccagatg gactaccgaa gattgcagcc ttcttgcctg
     2761 gggaccatgc ccctgtctgt gtgcccaccc atggctgagt cagggtcctg ctgtaccacc
     2821 cacattgcca accattccta cctaccacta agctattggc agcagccttg actcgagtct
     2881 agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     2941 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     3001 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     3061 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     3121 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     3181 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3241 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     3301 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     3361 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     3421 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     3481 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     3541 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     3601 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     3661 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     3721 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3781 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     3841 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     3901 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     3961 tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga ggatcgtttc
     4021 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     4081 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     4141 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     4201 tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     4261 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     4321 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     4381 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     4441 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     4501 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg
     4561 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     4621 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     4681 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     4741 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     4801 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     4861 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     4921 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     4981 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     5041 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     5101 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     5161 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     5221 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     5281 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     5341 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     5401 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     5461 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     5521 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     5581 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     5641 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     5701 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     5761 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     5821 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     5881 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     5941 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     6001 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     6061 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     6121 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     6181 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     6241 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     6301 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     6361 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     6421 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     6481 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     6541 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     6601 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     6661 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     6721 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     6781 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     6841 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     6901 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     6961 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     7021 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     7081 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     7141 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     7201 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     7261 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
