LOCUS       Exported                8166 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  expression of U2AF1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pRRL_U2AF1_WT_mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8166)
  AUTHORS   Ilagan JO, Ramakrishnan A, Hayes B, Murphy ME, Zebari AS, Bradley P,
            Bradley RK
  TITLE     U2AF1 mutations alter splice site recognition in hematological 
            malignancies.
  JOURNAL   Genome Res. 2015 Jan;25(1):14-26. doi: 10.1101/gr.181016.114. Epub 
            2014 Sep 29.
  PUBMED    25267526
REFERENCE   2  (bases 1 to 8166)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8166
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        27..131
                     /gene="bla"
                     /label=AmpR promoter
     CDS             132..992
                     /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     complement(350..369)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1163..1751
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1652..1671
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1905..1922
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2039..2060
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        2075..2105
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2113..2129
                     /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)."
     primer_bind     2118..2140
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2137..2153
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2137..2153
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2172..2192
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2174..2192
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     LTR             2448..2628
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    2672..2797
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    3290..3523
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             3707..3751
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    4018..4135
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        4181..4691
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     4610..4628
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             5429..5452
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             5453..5506
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             5507..6217
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(5652..5670)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5920..5939
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    6232..6820
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6285..6305)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6703..6714)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             6907..7140
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    7212..7333
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7249..7268)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7303..7322
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      7352..7487
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7414..7433
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(7519..7538)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(7520..7538)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7548..7565)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7548..7564)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7557..7579)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      7709..8164
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7796..7815)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8006..8027
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
       61 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
      121 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
      181 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
      241 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
      301 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
      361 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
      421 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
      481 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
      541 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
      601 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
      661 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
      721 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
      781 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
      841 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
      901 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
      961 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     1021 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     1081 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     1141 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     1201 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     1261 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     1321 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     1381 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     1441 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     1501 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     1561 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     1621 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     1681 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     1741 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     1801 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     1861 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     1921 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     1981 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     2041 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     2101 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
     2161 acgccaagcg cgcaattaac cctcactaaa gggaacaaaa gctggagctg caagcttaat
     2221 gtagtcttat gcaatactct tgtagtcttg caacatggta acgatgagtt agcaacatgc
     2281 cttacaagga gagaaaaagc accgtgcatg ccgattggtg gaagtaaggt ggtacgatcg
     2341 tgccttatta ggaaggcaac agacgggtct gacatggatt ggacgaacca ctgaattgcc
     2401 gcattgcaga gatattgtat ttaagtgcct agctcgatac aataaacggg tctctctggt
     2461 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
     2521 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
     2581 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa
     2641 cagggacctg aaagcgaaag ggaaaccaga gctctctcga cgcaggactc ggcttgctga
     2701 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag
     2761 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag
     2821 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
     2881 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
     2941 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
     3001 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
     3061 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
     3121 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt
     3181 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
     3241 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
     3301 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcctcaatg acgctgacgg
     3361 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
     3421 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
     3481 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
     3541 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
     3601 tggaacagat tggaatcaca cgacctggat ggagtgggac agagaaatta acaattacac
     3661 aagcttaata cactccttaa ttgaagaatc gcaaaaccag caagaaaaga atgaacaaga
     3721 attattggaa ttagataaat gggcaagttt gtggaattgg tttaacataa caaattggct
     3781 gtggtatata aaattattca taatgatagt aggaggcttg gtaggtttaa gaatagtttt
     3841 tgctgtactt tctatagtga atagagttag gcagggatat tcaccattat cgtttcagac
     3901 ccacctccca accccgaggg gacccgacag gcccgaagga atagaagaag aaggtggaga
     3961 gagagacaga gacagatcca ttcgattagt gaacggatct cgacggtatc ggttaacttt
     4021 taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag acataatagc
     4081 aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa attttccgat
     4141 cacgagacta gcctcgagaa gcttgatatc gaattcccac ggggttgggg ttgcgccttt
     4201 tccaaggcag ccctgggttt gcgcagggac gcggctgctc tgggcgtggt tccgggaaac
     4261 gcagcggcgc cgaccctggg tctcgcacat tcttcacgtc cgttcgcagc gtcacccgga
     4321 tcttcgccgc tacccttgtg ggccccccgg cgacgcttcc tgctccgccc ctaagtcggg
     4381 aaggttcctt gcggttcgcg gcgtgccgga cgtgacaaac ggaagccgca cgtctcacta
     4441 gtaccctcgc agacggacag cgccagggag caatggcagc gcgccgaccg cgatgggctg
     4501 tggccaatag cggctgctca gcggggcgcg ccgagagcag cggccgggaa ggggcggtgc
     4561 gggaggcggg gtgtggggcg gtagtgtggg ccctgttcct gcccgcgcgg tgttccgcat
     4621 tctgcaagcc tccggagcgc acgtcggcag tcggctccct cgttgaccga atcaccgacc
     4681 tctctcccca gggggatccg aaatggcgga gtatctggcc tccatcttcg gcaccgagaa
     4741 agacaaagtc aactgttcat tttatttcaa aattggagca tgtcgtcatg gagacaggtg
     4801 ctctcggttg cacaataaac cgacgtttag ccagaccatt gccctcttga acatttaccg
     4861 taaccctcaa aactcttccc agtctgctga cggtttgcgc tgtgccgtga gcgatgtgga
     4921 gatgcaggaa cactatgatg agttttttga ggaggttttt acagaaatgg aggagaagta
     4981 tggggaagta gaggagatga acgtctgtga caacctggga gaccacctgg tggggaacgt
     5041 gtacgtcaag tttcgccgtg aggaagatgc ggaaaaggct gtgattgact tgaataaccg
     5101 ttggtttaat ggacagccga tccacgccga gctgtcaccc gtgacggact tcagagaagc
     5161 ctgctgccgt cagtatgaga tgggagaatg cacacgaggc ggcttctgca acttcatgca
     5221 tttgaagccc atttccagag agctgcggcg ggagctgtat ggccgccgtc gcaagaagca
     5281 tagatcaaga tcccgatccc gggagcgtcg ttctcggtct agagaccgtg gtcgtggcgg
     5341 tggcggtggc ggtggtggag gtggcggcgg acgggagcgt gacaggaggc ggtcgagaga
     5401 tcgtgaaaga tctgggcgat tcggcggcga ctacaaagac gatgacgaca aggagggcag
     5461 aggaagtctt ctaacatgcg gtgacgtgga ggagaatccc ggccctatgg tgagcaaggg
     5521 cgaggaggat aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg
     5581 ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg
     5641 cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat
     5701 cctgtcccct cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc
     5761 cgactacttg aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga
     5821 ggacggcggc gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta
     5881 caaggtgaag ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac
     5941 catgggctgg gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga
     6001 gatcaagcag aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac
     6061 ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga
     6121 catcacctcc cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg
     6181 ccactccacc ggcggcatgg acgagctgta caagtaagcg gccgcctcga caatcaacct
     6241 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg
     6301 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc
     6361 attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt
     6421 gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc
     6481 attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg
     6541 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact
     6601 gacaattccg tggtgttgtc ggggaagctg acgtcctttc catggctgct cgcctgtgtt
     6661 gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg
     6721 gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc
     6781 cctcagacga gtcggatctc cctttgggcc gcctccccgc ctggaattcg agctcggtac
     6841 ctttaagacc aatgacttac aaggcagctg tagatcttag ccacttttta aaagaaaagg
     6901 ggggactgga agggctaatt cactcccaac gaagacaaga tctgcttttt gcttgtactg
     6961 ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac
     7021 tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt
     7081 gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca
     7141 gtagtagttc atgtcatctt attattcagt atttataact tgcaaagaaa tgaatatcag
     7201 agagtgagag gaacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     7261 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     7321 tcaatgtatc ttatcatgtc tggctctagc tatcccgccc ctaactccgc ccagttccgc
     7381 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     7441 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcgt
     7501 cgagacgtac ccaattcgcc ctatagtgag tcgtattacg cgcgctcact ggccgtcgtt
     7561 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
     7621 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     7681 ttgcgcagcc tgaatggcga atggcgcgac gcgccctgta gcggcgcatt aagcgcggcg
     7741 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     7801 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     7861 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     7921 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     7981 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     8041 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     8101 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca
     8161 atttcc
//
