LOCUS       sequence-335788-        7232 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of 3x-Flag-tagged human PP1 beta catalytic 
            subunit.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7232)
  TITLE     Balk lab plasmids
REFERENCE   2  (bases 1 to 7232)
  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"
COMMENT     Sequence Label: Flag-neo-PP1-Beta
FEATURES             Location/Qualifiers
     source          1..7232
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     126..148
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     140..157
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     141..157
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        318..697
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        698..901
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     852..872
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             931..996
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    1991..2613
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2190..2209)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2637..2657)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2655..2952
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             3006..3800
                     /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(3060..3079)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3670..3689
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4452..4586
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4489..4508)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4543..4562
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4591..4610)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4592..4610)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4624..4640)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4624..4640)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4637..4659)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4648..4664
                     /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(4672..4678,4679..4696,4697..4702))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4717..4738
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4855..4872)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5026..5614)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5106..5125)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5785..6576,6577..6645))
                     /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     6408..6427
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6646..6750)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6777..7205
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6864..6883)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7074..7095
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
       61 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
      121 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctgatct atacattgaa
      181 tcaatattgg caattagcca tattagtcat tggttatata gcataaatca atattggcta
      241 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      301 aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat caattacggg
      361 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      421 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      481 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      541 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      601 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      661 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      721 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      781 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      841 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      901 tcgtttagtg aaccgtcaga attaaccatg gactacaaag accatgacgg tgattataaa
      961 gatcatgaca tcgattacaa ggatgacgat gacaagctta tggcggacgg ggagctgaac
     1021 gtggacagcc tcatcacccg gctgctggag gtacgaggat gtcgtccagg aaagattgtg
     1081 cagatgactg aagcagaagt tcgaggctta tgtatcaagt ctcgggagat ctttctcagc
     1141 cagcctattc ttttggaatt ggaagcaccg ctgaaaattt gtggagatat tcatggacag
     1201 tatacagatt tactgagatt atttgaatat ggaggtttcc caccagaagc caactatctt
     1261 ttcttaggag attatgtgga cagaggaaag cagtctttgg aaaccatttg tttgctattg
     1321 gcttataaaa tcaaatatcc agagaacttc tttctcttaa gaggaaacca tgagtgtgct
     1381 agcatcaatc gcatttatgg attctatgat gaatgcaaac gaagatttaa tattaaattg
     1441 tggaagacct tcactgattg ttttaactgt ctgcctatag cagccattgt ggatgagaag
     1501 atcttctgtt gtcatggagg attgtcacca gacctgcaat ctatggagca gattcggaga
     1561 attatgagac ctactgatgt ccctgataca ggtttgctct gtgatttgct atggtctgat
     1621 ccagataagg atgtgcaagg ctggggagaa aatgatcgtg gtgtttcctt tacttttgga
     1681 gctgatgtag tcagtaaatt tctgaatcgt catgatttag atttgatttg tcgagctcat
     1741 caggtggtgg aagatggata tgaatttttt gctaaacgac agttggtaac cttattttca
     1801 gccccaaatt actgtggcga gtttgataat gctggtggaa tgatgagtgt ggatgaaact
     1861 ttgatgtgtt catttcagat attgaaacca tctgaaaaga aagctaaata ccagtatggt
     1921 ggactgaatt ctggacgtcc tgtcactcca cctcgaacag ctaatccgcc gaagaaaagg
     1981 tgaggatccc gggtggcatc cctgtgaccc ctccccagtg cctctcctgg ccctggaagt
     2041 tgccactcca gtgcccacca gccttgtcct aataaaatta agttgcatca ttttgtctga
     2101 ctaggtgtcc ttctataata ttatggggtg gaggggggtg gtatggagca aggggcaagt
     2161 tgggaagaca acctgtaggg cctgcggggt ctattgggaa ccaagctgga gtgcagtggc
     2221 acaatcttgg ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc tgcctcagcc
     2281 tcccgagttg ttgggattcc aggcatgcat gaccaggctc agctaatttt tgtttttttg
     2341 gtagagacgg ggtttcacca tattggccag gctggtctcc aactcctaat ctcaggtgat
     2401 ctacccacct tggcctccca aattgctggg attacaggcg tgaaccactg ctcccttccc
     2461 tgtccttctg attttaaaat aactatacca gcaggaggac gtccagacac agcataggct
     2521 acctggccat gcccaaccgg tgggacattt gagttgcttg cttggcactg tcctctcatg
     2581 cgttgggtcc actcagtaga tgcctgttga attgggtacg cggccagctt ggctgtggaa
     2641 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     2701 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     2761 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     2821 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     2881 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     2941 aggctttttt ggaggaattg atcagcttgg gatctgatca agagacagga tgaggatcgt
     3001 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     3061 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     3121 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     3181 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     3241 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     3301 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     3361 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     3421 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     3481 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc
     3541 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     3601 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     3661 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     3721 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     3781 ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
     3841 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     3901 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     3961 cttcgcccac cccgggctcg atcccctcgc gagttggttc agctgctgcc tgaggctgga
     4021 cgacctcgcg gagttctacc ggcagtgcaa atccgtcggc atccaggaaa ccagcagcgg
     4081 ctatccgcgc atccatgccc ccgaactgca ggagtgggga ggcacgatgg ccgctttggt
     4141 cgacccggac gggacgctcc tgcgcctgat acagaacgaa ttgcttgcag gcatctcatg
     4201 agtgtgtctt cccgttttcc gcctgaggtc actgcgtgga tggagcgctg gcgcctgctg
     4261 cgcgacggcg agctgctcac cacccactcg ccaagctgga accgtaaaaa ggccgcgttg
     4321 ctggcgtttt tccataggct ccgccgatca taatcagcca taccacattt gtagaggttt
     4381 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
     4441 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     4501 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     4561 tcaatgtatc ttatcatgtc tggatcaatt ccctatagtg agtcgtatta aattcgtaat
     4621 catgtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     4681 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     4741 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     4801 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     4861 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     4921 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     4981 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     5041 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     5101 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     5161 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     5221 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     5281 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     5341 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     5401 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     5461 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     5521 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     5581 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     5641 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     5701 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     5761 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     5821 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     5881 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     5941 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     6001 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     6061 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     6121 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     6181 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     6241 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     6301 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     6361 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     6421 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     6481 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     6541 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     6601 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     6661 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     6721 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgc
     6781 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     6841 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     6901 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     6961 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     7021 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     7081 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     7141 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     7201 gaattttaac aaaatattaa cgcttacaat tt
//
