LOCUS       Exported                8034 bp ds-DNA     circular SYN 23-SEP-2021
DEFINITION  Expresses PIK3CD in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    EGFP-PIK3CD
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8034)
  AUTHORS   Zhou Y, Feng Z, Cao F, Liu X, Xia X, Yu CH
  TITLE     Abl-mediated PI3K activation regulates macrophage podosome 
            formation.
  JOURNAL   J Cell Sci. 2020 Jun 8;133(11). pii: jcs.234385. doi: 
            10.1242/jcs.234385.
  PUBMED    32393599
REFERENCE   2  (bases 1 to 8034)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 23, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8034
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        49..1026
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1061..1079
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1061..1078
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      1145..1154
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1151..1867
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             1151..1867
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1196..1217)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1457..1476)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1804..1825
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    1889..1913
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(5049..5073)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     promoter        complement(5090..5107)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    5112..5246
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5136..5155)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5190..5209
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5359..5379)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5359..5377)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5398..5414)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5398..5414)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5411..5433)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5422..5438
                     /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(5446..5476)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5491..5512
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5629..5646)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5800..6388)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5880..5899)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6559..7419)
                     /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     7182..7201
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7420..7524)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(7550..8005)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7687..7708)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7899..7918
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agtcgaccgc caattcaata
       61 tggcgtatat ggactcatgc caattcaata tggtggatct ggacctgtgc caattcaata
      121 tggcgtatat ggactcgtgc caattcaata tggtggatct ggaccccagc caattcaata
      181 tggcggactt ggcaccatgc caattcaata tggcggacct ggcactgtgc caactgggga
      241 ggggtctact tggcacggtg ccaagtttga ggaggggtct tggccctgtg ccaagtccgc
      301 catattgaat tggcatggtg ccaataatgg cggccatatt ggctatatgc caggatcaat
      361 atataggcaa tatccaatat ggccctatgc caatatggct attggccagg ttcaatacta
      421 tgtattggcc ctatgccata tagtattcca tatatgggtt ttcctattga cgtagatagc
      481 ccctcccaat gggcggtccc atataccata tatggggctt cctaataccg cccatagcca
      541 ctcccccatt gacgtcaatg gtctctatat atggtctttc ctattgacgt catatgggcg
      601 gtcctattga cgtatatggc gcctccccca ttgacgtcaa ttacggtaaa tggcccgcct
      661 ggctcaatgc ccattgacgt caataggacc acccaccatt gacgtcaatg ggatggctca
      721 ttgcccattc atatccgttc tcacgccccc tattgacgtc aatgacggta aatggcccac
      781 ttggcagtac atcaatatct attaatagta acttggcaag tacattacta ttggaagtac
      841 gccagggtac attggcagta ctcccattga cgtcaatggc ggtaaatggc ccgcgatggc
      901 tgccaagtac atccccattg acgtcaatgg ggaggggcaa tgacgcaaat gggcgttcca
      961 ttgacgtaaa tgggcggtag gcgtgcctaa tgggaggtct atataagcaa tgctcgttta
     1021 gggaaccgcc attctgcctg gggacgtcgg agcaagcttg atttaggtga cactatagaa
     1081 tacaagctac ttgttctttt tgcaggatcc catcgattcg aattcaaggc tagcgctacc
     1141 ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     1201 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     1261 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     1321 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     1381 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     1441 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     1501 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     1561 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     1621 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     1681 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     1741 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     1801 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     1861 gtacaagtcc ggactcagat ctcgaatcac aagtttgtac aaaaaagttg gcaccatgcc
     1921 ccctggggtg gactgcccca tggaattctg gaccaaggag gagaatcaga gcgttgtggt
     1981 tgacttcctg ctgcccacag gggtctacct gaacttccct gtgtcccgca atgccaacct
     2041 cagcaccatc aagcagctgc tgtggcaccg cgcccagtat gagccgctct tccacatgct
     2101 cagtggcccc gaggcctatg tgttcacctg catcaaccag acagcggagc agcaagagct
     2161 ggaggacgag caacggcgtc tgtgtgacgt gcagcccttc ctgcccgtcc tgcgcctggt
     2221 ggcccgtgag ggcgaccgcg tgaagaagct catcaactca cagatcagcc tcctcatcgg
     2281 caaaggcctc cacgagtttg actccttgtg cgacccagaa gtgaacgact ttcgcgccaa
     2341 gatgtgccaa ttctgcgagg aggcggccgc ccgccggcag cagctgggct gggaggcctg
     2401 gctgcagtac agtttccccc tgcagctgga gccctcggct caaacctggg ggcctggtac
     2461 cctgcggctc ccgaaccggg cccttctggt caacgttaag tttgagggca gcgaggagag
     2521 cttcaccttc caggtgtcca ccaaggacgt gccgctggcg ctgatggcct gtgccctgcg
     2581 gaagaaggcc acagtgttcc ggcagccgct ggtggagcag ccggaagact acacgctgca
     2641 ggtgaacggc aggcatgagt acctgtatgg cagctacccg ctctgccagt tccagtacat
     2701 ctgcagctgc ctgcacagtg ggttgacccc tcacctgacc atggtccatt cctcctccat
     2761 cctcgccatg cgggatgagc agagcaaccc tgccccccag gtccagaaac cgcgtgccaa
     2821 accacctccc attcctgcga agaagccttc ctctgtgtcc ctgtggtccc tggagcagcc
     2881 gttccgcatc gagctcatcc agggcagcaa agtgaacgcc gacgagcgga tgaagctggt
     2941 ggtgcaggcc gggcttttcc acggcaacga gatgctgtgc aagacggtgt ccagctcgga
     3001 ggtgagcgtg tgctcggagc ccgtgtggaa gcagcggctg gagttcgaca tcaacatctg
     3061 cgacctgccc cgcatggccc gtctctgctt tgcgctgtac gccgtgatcg agaaagccaa
     3121 gaaggctcgc tccaccaaga agaagtccaa gaaggcggac tgccccattg cctgggccaa
     3181 cctcatgctg tttgactaca aggaccagct taagaccggg gaacgctgcc tctacatgtg
     3241 gccctccgtc ccagatgaga agggcgagct gctgaacccc acgggcactg tgcgcagtaa
     3301 ccccaacacg gatagcgccg ctgccctgct catctgcctg cccgaggtgg ccccgcaccc
     3361 cgtgtactac cccgccctgg agaagatctt ggagctgggg cgacacagcg agtgtgtgca
     3421 tgtcaccgag gaggagcagc tgcagctgcg ggaaatcctg gagcggcggg ggtctgggga
     3481 gctgtatgag cacgagaagg acctggtgtg gaagctgcgg catgaagtcc aggagcactt
     3541 cccggaggcg ctagcccggc tgctgctggt caccaagtgg aacaagcatg aggatgtggc
     3601 ccagatgctc tacctgctgt gctcctggcc ggagctgccc gtcctgagcg ccctggagct
     3661 gctagacttc agcttccccg attgccacgt aggctccttc gccatcaagt cgctgcggaa
     3721 actgacggac gatgagctgt tccagtacct gctgcagctg gtgcaggtgc tcaagtacga
     3781 gtcctacctg gactgcgagc tgaccaaatt cctgctggac cgggccctgg ccaaccgcaa
     3841 gatcggccac ttccttttct ggcacctccg ctccgagatg cacgtgccgt cggtggccct
     3901 gcgcttcggc ctcatcctgg aggcctactg caggggcagc acccaccaca tgaaggtgct
     3961 gatgaagcag ggggaagcac tgagcaaact gaaggccctg aatgacttcg tcaagctgag
     4021 ctctcagaag acccccaagc cccagaccaa ggagctgatg cacttgtgca tgcggcagga
     4081 ggcctaccta gaggccctct cccacctgca gtccccactc gaccccagca ccctgctggc
     4141 tgaagtctgc gtggagcagt gcaccttcat ggactccaag atgaagcccc tgtggatcat
     4201 gtacagcaac gaggaggcag gcagcggcgg cagcgtgggc atcatcttta agaacgggga
     4261 tgacctccgg caggacatgc tgaccctgca gatgatccag ctcatggacg tcctgtggaa
     4321 gcaggagggg ctggacctga ggatgacccc ctatggctgc ctccccaccg gggaccgcac
     4381 aggcctcatt gaggtggtac tccgttcaga caccatcgcc aacatccaac tcaacaagag
     4441 caacatggca gccacagccg ccttcaacaa ggatgccctg ctcaactggc tgaagtccaa
     4501 gaacccgggg gaggccctgg atcgagccat tgaggagttc accctctcct gtgctggcta
     4561 ttgtgtggcc acatatgtgc tgggcattgg cgatcggcac agcgacaaca tcatgatccg
     4621 agagagtggg cagctgttcc acattgattt tggccacttt ctggggaatt tcaagaccaa
     4681 gtttggaatc aaccgcgagc gtgtcccatt catcctcacc tacgactttg tccatgtgat
     4741 tcagcagggg aagactaata atagtgagaa atttgaacgg ttccggggct actgtgaaag
     4801 ggcctacacc atcctgcggc gccacgggct tctcttcctc cacctctttg ccctgatgcg
     4861 ggcggcaggc ctgcctgagc tcagctgctc caaagacatc cagtatctca aggactccct
     4921 ggcactgggg aaaacagagg aggaggcact gaagcacttc cgagtgaagt ttaacgaagc
     4981 cctccgtgag agctggaaaa ccaaagtgaa ctggctggcc cacaacgtgt ccaaagacaa
     5041 caggcagttg ccaactttct tgtacaaagt ggtgatcgag cctctagaac tatagtgagt
     5101 cgtattacgt agatccagac atgataagat acattgatga gtttggacaa accacaacta
     5161 gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa
     5221 ccattataag ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg
     5281 ttcaggggga ggtgtgggag gttttttaat tcgcggccgc ggcgccaatg cattgggccc
     5341 ggtacccagc ttttgttccc tttagtgagg gttaattgcg cgcttggcgt aatcatggtc
     5401 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     5461 gagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     5521 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     5581 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     5641 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     5701 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     5761 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     5821 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     5881 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     5941 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     6001 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     6061 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     6121 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     6181 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     6241 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     6301 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     6361 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     6421 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     6481 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     6541 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     6601 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     6661 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     6721 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     6781 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     6841 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     6901 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     6961 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     7021 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     7081 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     7141 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     7201 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     7261 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     7321 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     7381 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     7441 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     7501 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc
     7561 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     7621 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     7681 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     7741 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     7801 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     7861 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     7921 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     7981 cttaatgcgc cgctacaggg cgcgtcccat tcgccattca ggctgcgcaa ctgt
//
