LOCUS       Exported                5537 bp ds-DNA     circular SYN 15-APR-2017
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    pCRIS-PITChv2-dTAG-BSD (BRD4)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5537)
  AUTHORS   Justin Roberts
  TITLE     Direct Submission
  JOURNAL   Exported Saturday, Apr 15, 2017 from SnapGene 3.3.3
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5537
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      86..674
                     /direction=RIGHT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     primer_bind     944..960
                     /note="M13 fwd"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     protein_bind    1416..1432
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /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"
     misc_feature    1544..1566
                     /note="PITCh-gRNA Target"
                     /note="color: #800080"
     misc_feature    1567..1586
                     /note="Cterm 5' microhomology (BRD4)"
                     /note="color: #cc99ff"
     misc_feature    1587
                     /note="To correct frame of BRD4"
                     /note="color: #ff00ff"
     CDS             1588..1632
                     /codon_start=1
                     /note="(GGGGS)3"
                     /note="color: #008080"
                     /translation="GGGGSGGGGSGGGGS"
     CDS             1633..1953
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /note="FKBP-V"
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /note="color: #993366"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             1963..1989
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /note="HA"
                     /note="color: #cc99b2"
                     /translation="YPYDVPDYA"
     CDS             1993..2019
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /note="HA"
                     /note="color: #cc99b2"
                     /translation="YPYDVPDYA"
     CDS             2023..2079
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /note="P2A"
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /note="color: #cc99b2"
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2080..2475
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /note="BSD"
                     /note="confers resistance to blasticidin"
                     /note="color: #ccffcc"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    2479..2498
                     /note="Cterm 3' microhomology (BRD4)"
                     /note="color: #cc99ff"
     misc_feature    2499..2521
                     /note="PITCh-gRNA Target"
                     /note="color: #800080"
     polyA_signal    2558..2669
                     /note="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     promoter        2792..3121
                     /note="SV40 promoter"
                     /note="SV40 enhancer and early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     rep_origin      2972..3107
                     /note="SV40 ori"
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     CDS             3170..4195
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /note="HygR"
                     /note="confers resistance to hygromycin"
                     /note="color: #ccffcc"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    4325..4446
                     /note="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     CDS             complement(4499..5359)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 4499 .. 5290 / #ccffcc
                        2: 5291 .. 5359 / #ccffcc / signal sequence
                     Cleavage site after base 5290"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5360..5451)
                     /gene="bla"
                     /note="AmpR promoter"
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 ctcatgacca aaatccctta acgtgagtta cgcgcgcgtc gttccactga gcgtcagacc
       61 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
      121 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
      181 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
      241 tgtagccgta gttagcccac cacttcaaga actctgtagc accgcctaca tacctcgctc
      301 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
      361 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
      421 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
      481 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
      541 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
      601 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
      661 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
      721 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
      781 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
      841 gcgaggaagc ggaaggcgag agtagggaac tgccaggcat caaactaagc agaaggcccc
      901 tgacggatgg cctttttgcg tttctacaaa ctctttctgt gttgtaaaac gacggccagt
      961 cttaagctcg ggccccctgg gcggttctga taacgagtaa tcgttaatcc gcaaataacg
     1021 taaaaacccg cttcggcggg tttttttatg gggagagttt agggaaagag catttgtcag
     1081 aatatttaag ggcgcctgtc actttgcttg atatatgaga attatttaac cttataaatg
     1141 agaaaaaagc aacgcacttt aaataagata cgttgctttt tcgattgatg aacacctata
     1201 attaaactat tcatctatta tttatgattt tttgtatata caatatttct agtttgttaa
     1261 agagaattaa gaaaataaat ctcgaaaata ataaagggaa aatcagtttt tgatatcaaa
     1321 attatacatg tcaacgataa tacaaaatat aatacaaact ataagatgtt atcagtattt
     1381 attatcattt agaataaatt ttgtgtcgcc cttaattgtg agcggataac aattacgagc
     1441 ttcatgcaca gtggcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg
     1501 ggtcattagt tcatagccca tatatggagt tccgcgttac atagcatcgt acgcgtacgt
     1561 gtttggattg acatgaattt ccagagtggt ggcggtggct cgggcggtgg tgggtcgggt
     1621 ggcggcggat ctggagtgca ggtggaaacc atctccccag gagacgggcg caccttcccc
     1681 aagcgcggcc agacctgcgt ggtgcactac accgggatgc ttgaagatgg aaagaaagtt
     1741 gattcctccc gggacagaaa caagcccttt aagtttatgc taggcaagca ggaggtgatc
     1801 cgaggctggg aagaaggggt tgcccagatg agtgtgggtc agagagccaa actgactata
     1861 tctccagatt atgcctatgg tgccactggg cacccaggca tcatcccacc acatgccact
     1921 ctcgtcttcg atgtggagct tctaaaactg gaaggctcgg gctaccccta cgacgtgccc
     1981 gactacgccg gctatccgta tgatgtcccg gactatgcag gcgctactaa cttcagcctg
     2041 ctgaagcagg ctggagacgt ggaggagaac cctggaccta tggccaagcc tttgtctcaa
     2101 gaagaatcca ccctcattga aagagcaacg gctacaatca acagcatccc catctctgaa
     2161 gactacagcg tcgccagcgc agctctctct agcgacggcc gcatcttcac tggtgtcaat
     2221 gtatatcatt ttactggggg accttgtgca gaactcgtgg tgctgggcac tgctgctgct
     2281 gcggcagctg gcaacctgac ttgtatcgtc gcgatcggaa atgagaacag gggcatcttg
     2341 agcccctgcg gacggtgccg acaggtgctt ctcgatctgc atcctgggat caaagccata
     2401 gtgaaggaca gtgatggaca gccgacggca gttgggattc gtgaattgct gccctctggt
     2461 tatgtgtggg agggctgatg atctattgtc aatatttgcc aaacacgtac gcgtacgatg
     2521 ctctagaatg ctgatgggct agcaaaatca gcctcgactg tgccttctag ttgccagcca
     2581 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     2641 ctttcctaat aaaatgagga aattgcatca caacactcaa ccctatctcg gtctattctt
     2701 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     2761 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     2821 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     2881 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2941 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     3001 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     3061 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     3121 aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtga tgaaaaagcc
     3181 tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtttccga
     3241 cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg
     3301 tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta
     3361 tcggcacttt gcatcggccg cgctcccgat tccggaagtg cttgacattg gggaattcag
     3421 cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc
     3481 tgaaaccgaa ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc
     3541 ggccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata
     3601 cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc actggcaaac
     3661 tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg
     3721 ggccgaggac tgccccgaag tccggcacct cgtgcacgcg gatttcggct ccaacaatgt
     3781 cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga tgttcgggga
     3841 ttcccaatac gaggtcgcca acatcttctt ctggaggccg tggttggctt gtatggagca
     3901 gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc ggctccgggc
     3961 gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg gcaatttcga
     4021 tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt
     4081 cgggcgtaca caaatcgccc gcagaagcgc ggccgtctgg accgatggct gtgtagaagt
     4141 actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg aatagcacgt
     4201 gctacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     4261 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     4321 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     4381 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     4441 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcatt
     4501 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     4561 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     4621 gcgctgcgat gataccgcga gaaccacgct caccggctcc ggatttatca gcaataaacc
     4681 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     4741 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     4801 ttgttgccat cgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     4861 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     4921 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     4981 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     5041 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     5101 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     5161 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     5221 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     5281 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     5341 ggaaatgttg aatactcata ttcttccttt ttcaatatta ttgaagcatt tatcagggtt
     5401 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggtca
     5461 gtgttacaac caattaacca attctgaaca ttatcgcgag cccatttata cctgaatatg
     5521 gctcataaca ccccttg
//
