LOCUS       pcDNA3-Endo-INPP        7725 bp    DNA     circular SYN 02-JUN-2025
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7725)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 2, 2025 from SnapGene 8.1.0
            https://www.snapgene.com
COMMENT     Imported using the Genbank importer. File name: 
            pcDNA3-EKAR4-2xFYVE.gb Serial Cloner Genbank Format 
            SerialCloner_Type=Degen. DNA SerialCloner_Comments= 
            SerialCloner_Ends=0,0,,0,
FEATURES             Location/Qualifiers
     source          1..7725
                     /mol_type="other DNA"
                     /note="color: #ffffff"
                     /organism="synthetic DNA construct"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
                     /note="color: #ffffff"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /note="color: #ffffff; direction: RIGHT"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     CDS             922..939
                     /codon_start=1
                     /label=6His
                     /note="color: #008000"
                     /translation="HHHHHH"
     CDS             943..975
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /note="color: #cc99b2"
                     /translation="MASMTGGQQMG"
     CDS             join(979..987,988..1002)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /note="This feature has 2 segments:
                        1:  979 ..  987 / #cc99b2
                        2:  988 .. 1002 / #cc99b2 / enterokinase site
                     Cleavage site after base 1002"
                     /translation="DLYDDDDK"
     CDS             1009..2301
                     /codon_start=1
                     /label=INPP5E(214-644)
                     /note="color: #005493"
                     /translation="SDLADYKLRAQPLLVRAHSSLGPGRPRSPLACDDCSLRSAKSSFS
                     LLAPIRSKDVRSRSYLEGSLLASGALLGADELARYFPDRNVALFVATWNMQGQKELPPS
                     LDEFLLPAEADYAQDLYVIGVQEGCSDRREWETRLQETLGPHYVLLSSAAHGVLYMSLF
                     IRRDLIWFCSEVECSTVTTRIVSQIKTKGALGISFTFFGTSFLFITSHFTSGDGKVAER
                     LLDYTRTVQALVLPRNVPDTNPYRSSAADVTTRFDEVFWFGDFNFRLSGGRTVVDALLC
                     QGLVVDVPALLQHDQLIREMRKGSIFKGFQEPDIHFLPSYKFDIGKDTYDSTSKQRTPS
                     YTDRVLYRSRHKGDICPVSYSSCPGIKTSDHRPVYGLFRVKVRPGRDNIPLAAGKFDRE
                     LYLLGIKRRISKEIQRQQALQSQNSSTIASVS"
     CDS             2314..3021
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /note="color: #ff0000"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     CDS             3031..3261
                     /codon_start=1
                     /label=FYVE Tag
                     /note="color: #800080"
                     /translation="ESDAMFAAERAPDWVDAEECHRCRVQFGVVTRKHHCRACGQIFCG
                     KCSSKYSTIPKFGIEKEVRVCEPCYEQLNKKA"
     polyA_signal    3322..3546
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     rep_origin      3592..4020
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        4034..4364
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     rep_origin      4215..4350
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     CDS             4431..5225
                     /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))"
                     /note="color: #ccffcc"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    5399..5520
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(5569..5585)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    5593..5609
                     /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)."
                     /note="color: #31849b"
     promoter        complement(join(5617..5623,5624..5641,5642..5647))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 5617 .. 5623 / #ffffff / -10
                        2: 5624 .. 5641 / #ffffff
                        3: 5642 .. 5647 / #ffffff / -35"
     protein_bind    5662..5683
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /note="color: #31849b"
     rep_origin      complement(5971..6559)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(join(6730..7521,7522..7590))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 6730 .. 7521 / #ccffcc
                        2: 7522 .. 7590 / #ccffcc / signal sequence
                     Cleavage site after base 7521"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(7591..7695)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct 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 atgggtggac 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 gcttgcggcc
      901 gccaccatgg tgcggggttc tcatcatcat catcatcatg gtatggctag catgactggt
      961 ggacagcaaa tgggtcggga tctgtacgac gatgacgata aggatccctc ggatcttgca
     1021 gactacaagc tccgcgcgca gccgctcctg gtgcgggccc acagcagcct gggccccggc
     1081 cggccgcgga gccccctggc ctgcgacgac tgttcccttc gctcggccaa atcctccttc
     1141 agcctcctgg cgcccatccg cagcaaggac gtccgcagca ggagttacct ggagggcagc
     1201 ctcctggcca gcggggccct gttgggggcg gatgagctgg cccgctactt cccagaccgg
     1261 aacgtggcac tcttcgtggc cacctggaac atgcagggcc agaaggagct cccgcccagc
     1321 ctggacgagt tcctgctccc agccgaggcc gactatgccc aggacctgta tgtcatcggg
     1381 gtccaggagg gctgttctga caggcgggag tgggagactc gtctgcagga gacgctgggc
     1441 ccgcactatg tgctgctgtc ctcggcggcc cacggcgtgc tctacatgtc gctcttcatc
     1501 cgcagggacc tcatctggtt ctgctcagag gtggagtgct ccacggtgac cacacgcatc
     1561 gtgtctcaga tcaagaccaa gggggccttg ggcatcagct tcaccttttt tggcacttcc
     1621 ttcctcttca tcacgtccca cttcacctca ggtgacggga aggtggcgga gcggctgctg
     1681 gactacacca ggactgtaca agccctggtc ctgcccagaa atgtgcccga caccaacccc
     1741 tatcgctcca gcgcagcgga cgtcaccacc cgcttcgatg aggtgttctg gtttggagac
     1801 ttcaacttcc gcctgagtgg cgggcgcaca gtcgtggacg ccctcctgtg ccagggcctg
     1861 gtggtggacg tgccggcgct gctgcagcac gaccagctca tccgggagat gcggaaaggg
     1921 tccatcttca agggcttcca ggagccggac atccacttcc tcccatcata caagtttgac
     1981 atcgggaagg acacgtacga cagcacctcc aagcagagga cgccctcata cacggaccgc
     2041 gtcttgtaca gaagccgcca caagggtgac atctgtcctg tgagctactc ttcctgcccc
     2101 gggatcaaga cgtccgacca ccgccctgtg tatggcctct tccgggtgaa agtgaggccg
     2161 gggcgagaca acattccgtt ggcagctggc aaatttgata gagaactgta cttactagga
     2221 attaaaagac ggatttcgaa ggagattcag aggcagcaag cactacagag tcagaactcc
     2281 agcaccatcg cctccgtttc tgctagcgat cccatggtga gcaagggcga ggaggataac
     2341 atggccatca tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc
     2401 cacgagttcg agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc
     2461 aagctgaagg tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag
     2521 ttcatgtacg gctccaaggc ctacgtgaag caccccgccg acatccccga ctacttgaag
     2581 ctgtccttcc ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgtg
     2641 gtgaccgtga cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg
     2701 cgcggcacca acttcccctc cgacggcccc gtaatgcaga agaagaccat gggctgggag
     2761 gcctcctccg agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg
     2821 ctgaagctga aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag
     2881 aagcccgtgc agctgcccgg cgcctacaac gtcaacatca agttggacat cacctcccac
     2941 aacgaggact acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc
     3001 ggcatggacg agctgtacaa gaaggaattc gaaagtgatg ccatgttcgc tgctgaaaga
     3061 gcccctgact gggtggatgc tgaggaatgc catcggtgca gagtacagtt tggggtggtg
     3121 acccgcaagc atcactgccg agcatgtggg cagatcttct gtggcaagtg ctcctccaag
     3181 tactccacca tccccaagtt cggcattgag aaggaggtgc gcgtgtgtga gccctgctat
     3241 gagcagctga acaagaaggc ataatctaga gggccctatt ctatagttca ctaaatgcta
     3301 gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     3361 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     3421 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     3481 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct
     3541 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct
     3601 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     3661 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     3721 gctttccccg tcaagctcta aatcggggca tccctttagg gttccgattt agtgctttac
     3781 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     3841 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     3901 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     3961 tggggatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     4021 aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc caggcaggca
     4081 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     4141 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     4201 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     4261 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     4321 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     4381 gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac
     4441 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact
     4501 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc
     4561 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg
     4621 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg
     4681 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt
     4741 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc
     4801 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag
     4861 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg
     4921 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc
     4981 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt
     5041 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg
     5101 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt
     5161 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct
     5221 tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg
     5281 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     5341 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     5401 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     5461 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     5521 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     5581 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     5641 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     5701 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     5761 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5821 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5881 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5941 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     6001 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     6061 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     6121 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcaatgct cacgctgtag
     6181 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     6241 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     6301 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     6361 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt
     6421 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     6481 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6541 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6601 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     6661 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     6721 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6781 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6841 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     6901 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6961 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     7021 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     7081 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     7141 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     7201 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     7261 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     7321 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     7381 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     7441 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     7501 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7561 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     7621 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     7681 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgt
//
