LOCUS       sequence-405477-        8611 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Single reporter randomly selected from splicing library for 
            biochemical validation..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8611)
  AUTHORS   Liao SE, Sudarshan M, Regev O
  TITLE     Deciphering RNA splicing logic with interpretable machine learning.
  JOURNAL   Proc Natl Acad Sci U S A. 2023 Oct 10;120(41):e2221165120. doi: 
            10.1073/pnas.2221165120. Epub 2023 Oct 5.
  PUBMED    37796983
REFERENCE   2  (bases 1 to 8611)
  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"; journalName: "Proc Natl 
            Acad Sci U S A."; date: "2023-10-10"; volume: "120(41):e2221165120. 
            doi"; pages: " 10.1073/pnas.2221165120. Epub 2023 Oct 5"
COMMENT     Sequence Label: pRegev_V2
FEATURES             Location/Qualifiers
     source          1..8611
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1112..1134
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1161..1180
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1161..1179
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2545..2562)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2551..2775
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2821..3249
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2908..2927)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3118..3139
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3258..3278)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3263..3592
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3443..3578
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3505..3524
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3640..3687
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3706..4080
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    4210..4331
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4247..4266)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4301..4320
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4380..4396)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4380..4396)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4393..4415)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4404..4420
                     /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(4428..4434,4435..4452,4453..4458))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4473..4494
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4611..4628)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4782..5370)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4862..4881)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5541..6332,6333..6401))
                     /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     6164..6183
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6402..6506)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(join(6569..7282,7283..7285,7286..7288))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(6614..6635)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     6963..6982
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     7222..7243
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      7285..7294
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(7296..8474)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          7305..8243
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(7328..7348)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     promoter        join(8505..8611,1..171)
                     /label=chicken beta-actin promoter
ORIGIN
        1 gggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg gggcggggcg
       61 aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt tccttttatg
      121 gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc gggagtcgct
      181 gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc gccccggctc
      241 tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc tccgggctgt
      301 aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga aagccttgag
      361 gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg cgtgtgtgtg
      421 tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg ctgcgggcgc
      481 ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg gggcggtgcc
      541 ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg
      601 gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca cccccctccc
      661 cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg tggcgcgggg
      721 ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc ggggccgcct
      781 cgggccgggg agggctcggg ggaggggcgc ggcggccccc ggagcgccgg cggctgtcga
      841 ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc gcagggactt
      901 cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac cccctctagc
      961 gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga gggccttcgt
     1021 gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac
     1081 ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt gaccggcggc
     1141 tctagcgttt aaacttaagc taatacgact cactataggg agcttggtac cgcaacctca
     1201 aacagacacc atggtgcacc tgactcctga ggagaagtct gccgttactg ccctgtgggg
     1261 caaggtgaac gtggatgaag ttggtggtga ggccctgggc aggttggtat caaggttaca
     1321 agacaggttt aaggagacca atagaaactg ggcatatgga gacagagaag actcttgggt
     1381 ttctgatagg cactgactct ctctgcctat gtctttctct gccatccagg tttcgcaccg
     1441 caggttcggt tttttggtat aatacgcggc actccaaaat tactctgggg caccctcacc
     1501 agcaggtctg actatgggac ccttgatgtt ttctttcccc ttcttttcta tggttaagtt
     1561 catgtcatag gaaggggaga agtaacaggg tacagtttag aatgggaaac agacgaatga
     1621 ttgcatcagt gtggaagtct caggatcgtt ttagtttctt ttatttgctg ttcataacaa
     1681 ttgttttctt ttgtttaatt cttgctttct ttttttttct tctccgcaat ttttactatt
     1741 atacttaatg ccttaacatt gtgtataaca aaaggaaata tctctgagat acattaagta
     1801 acttaaaaaa aaactttaca cagtctgcct agtacattac tatttggaat atatgtgtgc
     1861 ttatttgcat attcataatc tccctacttt attttctttt atttttaatt gatacataat
     1921 cattatacat atttatgggt taaagtgtaa tgttttaata tcgatacaca tattgaccaa
     1981 atcagggtaa ttttgcattt gtaattttaa aaaatgcttt cttcttttaa tatacttttt
     2041 tgtttatctt atttctaata ctttccctaa tctctttctt tcagggcaat aatgatacaa
     2101 tgtatcatgc ctctttgcac cattctaaag aataacagtg ataatttctg ggttaaggca
     2161 atagcaatat ttctgcatat aaatatttct gcatataaat tgtaactgat gtaagaggtt
     2221 tcatattgct aatagcagct acaatccagc taccattctg cttttatttt atggttggga
     2281 taaggctgga ttattctgag tccaagctag gcccttttgc taatcatgtt catacctctt
     2341 atcttcctcc cacagctcct gggcaacgtg ctggtctgtg tgctggccca tcactttggc
     2401 aaagaattca ccccaccagt gcaggctgcc tatcagaaag tggtggctgg tgtggctaat
     2461 gccctggccc acaagtatca ctaagctcgc tctagaatac cggtagtgtg tctattatag
     2521 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
     2581 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     2641 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     2701 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
     2761 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
     2821 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     2881 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     2941 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     3001 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     3061 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     3121 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     3181 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     3241 acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     3301 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     3361 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     3421 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     3481 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga
     3541 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc
     3601 gggagcttgt atatccattt tcggatctga tcagcacgtg ttgacaatta atcatcggca
     3661 tagtatatcg gcatagtata atacgacaag gtgaggaact aaaccatggc caagttgacc
     3721 agtgccgttc cggtgctcac cgcgcgcgac gtcgccggag cggtcgagtt ctggaccgac
     3781 cggctcgggt tctcccggga cttcgtggag gacgacttcg ccggtgtggt ccgggacgac
     3841 gtgaccctgt tcatcagcgc ggtccaggac caggtggtgc cggacaacac cctggcctgg
     3901 gtgtgggtgc gcggcctgga cgagctgtac gccgagtggt cggaggtcgt gtccacgaac
     3961 ttccgggacg cctccgggcc ggccatgacc gagatcggcg agcagccgtg ggggcgggag
     4021 ttcgccctgc gcgacccggc cggcaactgc gtgcacttcg tggccgagga gcaggactga
     4081 cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     4141 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     4201 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     4261 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     4321 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     4381 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     4441 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     4501 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4561 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     4621 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     4681 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     4741 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     4801 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     4861 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     4921 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     4981 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     5041 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5101 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5161 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5221 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5281 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     5341 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     5401 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5461 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     5521 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     5581 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     5641 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     5701 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     5761 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     5821 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     5881 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     5941 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6001 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     6061 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     6121 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     6181 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     6241 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     6301 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6361 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6421 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6481 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac
     6541 ggatcgggag atcccagtgt ggtggtactt acttgtacag ctcgtccatg ccgagagtga
     6601 tcccggcggc ggtcacgaac tccagcagga ccatgtgatc gcgcttctcg ttggggtctt
     6661 tgctcagggc ggactgggtg ctcaggtagt ggttgtcggg cagcagcacg gggccgtcgc
     6721 cgatgggggt gttctgctgg tagtggtcgg cgagctgcac gctgccgtcc tcgatgttgt
     6781 ggcggatctt gaagttcgcc ttgatgccgt tcttctgctt gtcggccatg atatagacgt
     6841 tgtggctgtt gtagttgtac tccagcttgt gccccaggat gttgccgtcc tccttgaagt
     6901 cgatgccctt cagctcgatg cggttcacca gggtgtcgcc ctcgaacttc acctcggcgc
     6961 gggtcttgta gttgccgtcg tccttgaaga agatggtgcg ctcctggacg tagccttcgg
     7021 gcatggcgga cttgaagaag tcgtgctgct tcatgtggtc ggggtagcgg ctgaagcact
     7081 gcacgccgta ggtcagggtg gtcacgaggg tgggccaggg cacgggcagc ttgccggtgg
     7141 tgcagatgaa cttcagggtc agcttgccgt aggtggcatc gccctcgccc tcgccagaca
     7201 cgctgaactt gtggccgttt acgtcgccgt ccagctcgac caggatgggc accaccccgg
     7261 tgaacagctc ctcgcccttg ctcaccatgg tggcctcacg acacctgaaa tggaagaaaa
     7321 aaactttgaa ccactgtctg aggcttgaga atgaaccaag atccaaactc aaaaagggca
     7381 aattccaagg agaattacat caagtgccaa gctggcctaa cttcagtctc cacccactca
     7441 gtgtggggaa actccatcgc ataaaacccc tccccccaac ctaaagacga cgtactccaa
     7501 aagctcgaga actaatcgag gtgcctggac ggcgcccggt actccgtgga gtcacatgaa
     7561 gcgacggctg aggacggaaa ggcccttttc ctttgtgtgg gtgactcacc cgcccgctct
     7621 cccgagcgcc gcgtcctcca ttttgagctc cctgcagcag ggccgggaag cggccatctt
     7681 tccgctcacg caactggtgc cgaccgggcc agccttgccg cccagggcgg ggcgatacac
     7741 ggcggcgcga ggccaggcac cagagcaggc cggccagctt gagactaccc ccgtccgatt
     7801 ctcggtggcc gcgctcgcag gccccgcctc gccgaacatg tgcgctggga cgcacgggcc
     7861 ccgtcgccgc ccgcggcccc aaaaaccgaa ataccagtgt gcagatcttg gcccgcattt
     7921 acaagactat cttgccagaa aaaaagcgtc gcagcaggtc atcaaaaatt ttaaatggct
     7981 agagacttat cgaaagcagc gagacaggcg cgaaggtgcc accagattcg cacgcggcgg
     8041 ccccagcgcc caggccaggc ctcaactcaa gcacgaggcg aaggggctcc ttaagcgcaa
     8101 ggcctcgaac tctcccaccc acttccaacc cgaagctcgg gatcaagaat cacgtactgc
     8161 agccaggtgg aagtaattca aggcacgcaa gggccataac ccgtaaagag gccaggcccg
     8221 cgggaaccac acacggcact tacctgtgtt ctggcggcaa acccgttgcg aaaaagaacg
     8281 ttcacggcga ctactgcact tatatacggt tctcccccac cctcgggaaa aaggcggagc
     8341 cagtacacga catcactttc ccagtttacc ccgcgccacc ttctctaggc accggttcaa
     8401 ttgccgaccc ctccccccaa cttctcgggg actgtgggcg atgtgcgctc tgcccactga
     8461 cgggcaccgg agcctacgta ggaattaatt cgccagcaca gtggtcgagg tgagccccac
     8521 gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat
     8581 tttttaatta ttttgtgcag cgatgggggc g
//
