LOCUS       sequence-405421-        8586 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Backbone for cloning individual exon sequences into beta globin 
            minigene splicing reporter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8586)
  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 8586)
  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_ExonSkip_LandingPad
FEATURES             Location/Qualifiers
     source          1..8586
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(44..757,758..760,761..763))
                     /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(89..110)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     438..457
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     697..718
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      760..769
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(771..1949)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          780..1718
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(803..823)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     promoter        1980..2257
                     /label=chicken beta-actin promoter
     primer_bind     3198..3220
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     3247..3266
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3247..3265
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4606..4623)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4612..4836
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4882..5310
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4969..4988)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5179..5200
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5319..5339)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5324..5653
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5504..5639
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5566..5585
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5701..5748
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5767..6141
                     /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    6271..6392
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6308..6327)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6362..6381
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6441..6457)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6441..6457)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6454..6476)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6465..6481
                     /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(6489..6495,6496..6513,6514..6519))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6534..6555
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6672..6689)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6843..7431)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6923..6942)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7602..8393,8394..8462))
                     /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     8225..8244
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8463..8567)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccacctgacg tcgacggatc gggagatccc agtgtggtgg tacttacttg tacagctcgt
       61 ccatgccgag agtgatcccg gcggcggtca cgaactccag caggaccatg tgatcgcgct
      121 tctcgttggg gtctttgctc agggcggact gggtgctcag gtagtggttg tcgggcagca
      181 gcacggggcc gtcgccgatg ggggtgttct gctggtagtg gtcggcgagc tgcacgctgc
      241 cgtcctcgat gttgtggcgg atcttgaagt tcgccttgat gccgttcttc tgcttgtcgg
      301 ccatgatata gacgttgtgg ctgttgtagt tgtactccag cttgtgcccc aggatgttgc
      361 cgtcctcctt gaagtcgatg cccttcagct cgatgcggtt caccagggtg tcgccctcga
      421 acttcacctc ggcgcgggtc ttgtagttgc cgtcgtcctt gaagaagatg gtgcgctcct
      481 ggacgtagcc ttcgggcatg gcggacttga agaagtcgtg ctgcttcatg tggtcggggt
      541 agcggctgaa gcactgcacg ccgtaggtca gggtggtcac gagggtgggc cagggcacgg
      601 gcagcttgcc ggtggtgcag atgaacttca gggtcagctt gccgtaggtg gcatcgccct
      661 cgccctcgcc agacacgctg aacttgtggc cgtttacgtc gccgtccagc tcgaccagga
      721 tgggcaccac cccggtgaac agctcctcgc ccttgctcac catggtggcc tcacgacacc
      781 tgaaatggaa gaaaaaaact ttgaaccact gtctgaggct tgagaatgaa ccaagatcca
      841 aactcaaaaa gggcaaattc caaggagaat tacatcaagt gccaagctgg cctaacttca
      901 gtctccaccc actcagtgtg gggaaactcc atcgcataaa acccctcccc ccaacctaaa
      961 gacgacgtac tccaaaagct cgagaactaa tcgaggtgcc tggacggcgc ccggtactcc
     1021 gtggagtcac atgaagcgac ggctgaggac ggaaaggccc ttttcctttg tgtgggtgac
     1081 tcacccgccc gctctcccga gcgccgcgtc ctccattttg agctccctgc agcagggccg
     1141 ggaagcggcc atctttccgc tcacgcaact ggtgccgacc gggccagcct tgccgcccag
     1201 ggcggggcga tacacggcgg cgcgaggcca ggcaccagag caggccggcc agcttgagac
     1261 tacccccgtc cgattctcgg tggccgcgct cgcaggcccc gcctcgccga acatgtgcgc
     1321 tgggacgcac gggccccgtc gccgcccgcg gccccaaaaa ccgaaatacc agtgtgcaga
     1381 tcttggcccg catttacaag actatcttgc cagaaaaaaa gcgtcgcagc aggtcatcaa
     1441 aaattttaaa tggctagaga cttatcgaaa gcagcgagac aggcgcgaag gtgccaccag
     1501 attcgcacgc ggcggcccca gcgcccaggc caggcctcaa ctcaagcacg aggcgaaggg
     1561 gctccttaag cgcaaggcct cgaactctcc cacccacttc caacccgaag ctcgggatca
     1621 agaatcacgt actgcagcca ggtggaagta attcaaggca cgcaagggcc ataacccgta
     1681 aagaggccag gcccgcggga accacacacg gcacttacct gtgttctggc ggcaaacccg
     1741 ttgcgaaaaa gaacgttcac ggcgactact gcacttatat acggttctcc cccaccctcg
     1801 ggaaaaaggc ggagccagta cacgacatca ctttcccagt ttaccccgcg ccaccttctc
     1861 taggcaccgg ttcaattgcc gacccctccc cccaacttct cggggactgt gggcgatgtg
     1921 cgctctgccc actgacgggc accggagcct acgtaggaat taattcgcca gcacagtggt
     1981 cgaggtgagc cccacgttct gcttcactct ccccatctcc cccccctccc cacccccaat
     2041 tttgtattta tttatttttt aattattttg tgcagcgatg ggggcggggg gggggggggg
     2101 gcgcgcgcca ggcggggcgg ggcggggcga ggggcggggc ggggcgaggc ggagaggtgc
     2161 ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct tttatggcga ggcggcggcg
     2221 gcggcggccc tataaaaagc gaagcgcgcg gcgggcggga gtcgctgcgc gctgccttcg
     2281 ccccgtgccc cgctccgccg ccgcctcgcg ccgcccgccc cggctctgac tgaccgcgtt
     2341 actcccacag gtgagcgggc gggacggccc ttctcctccg ggctgtaatt agcgcttggt
     2401 ttaatgacgg cttgtttctt ttctgtggct gcgtgaaagc cttgaggggc tccgggaggg
     2461 ccctttgtgc ggggggagcg gctcgggggg tgcgtgcgtg tgtgtgtgcg tggggagcgc
     2521 cgcgtgcggc tccgcgctgc ccggcggctg tgagcgctgc gggcgcggcg cggggctttg
     2581 tgcgctccgc agtgtgcgcg aggggagcgc ggccgggggc ggtgccccgc ggtgcggggg
     2641 gggctgcgag gggaacaaag gctgcgtgcg gggtgtgtgc gtgggggggt gagcaggggg
     2701 tgtgggcgcg tcggtcgggc tgcaaccccc cctgcacccc cctccccgag ttgctgagca
     2761 cggcccggct tcgggtgcgg ggctccgtac ggggcgtggc gcggggctcg ccgtgccggg
     2821 cggggggtgg cggcaggtgg gggtgccggg cggggcgggg ccgcctcggg ccggggaggg
     2881 ctcgggggag gggcgcggcg gcccccggag cgccggcggc tgtcgaggcg cggcgagccg
     2941 cagccattgc cttttatggt aatcgtgcga gagggcgcag ggacttcctt tgtcccaaat
     3001 ctgtgcggag ccgaaatctg ggaggcgccg ccgcaccccc tctagcgggc gcggggcgaa
     3061 gcggtgcggc gccggcagga aggaaatggg cggggagggc cttcgtgcgt cgccgcgccg
     3121 ccgtcccctt ctccctctcc agcctcgggg ctgtccgcgg ggggacggct gccttcgggg
     3181 gggacggggc agggcggggt tcggcttctg gcgtgtgacc ggcggctcta gcgtttaaac
     3241 ttaagctaat acgactcact atagggagct tggtaccgca acctcaaaca gacaccatgg
     3301 tgcacctgac tcctgaggag aagtctgccg ttactgccct gtggggcaag gtgaacgtgg
     3361 atgaagttgg tggtgaggcc ctgggcaggt tggtatcaag gttacaagac aggtttaagg
     3421 agaccaatag aaactgggca tatggagaca gagaagactc ttgggtttct gataggcact
     3481 gactctctct gcctatgtct ttctctgcca tccaggttga gacgtaatcg agcgcaaagc
     3541 ggccgcaaat cgcgttgcgt ctccaggtct gactatggga cccttgatgt tttctttccc
     3601 cttcttttct atggttaagt tcatgtcata ggaaggggag aagtaacagg gtacagttta
     3661 gaatgggaaa cagacgaatg attgcatcag tgtggaagtc tcaggatcgt tttagtttct
     3721 tttatttgct gttcataaca attgttttct tttgtttaat tcttgctttc tttttttttc
     3781 ttctccgcaa tttttactat tatacttaat gccttaacat tgtgtataac aaaaggaaat
     3841 atctctgaga tacattaagt aacttaaaaa aaaactttac acagtctgcc tagtacatta
     3901 ctatttggaa tatatgtgtg cttatttgca tattcataat ctccctactt tattttcttt
     3961 tatttttaat tgatacataa tcattataca tatttatggg ttaaagtgta atgttttaat
     4021 atcgatacac atattgacca aatcagggta attttgcatt tgtaatttta aaaaatgctt
     4081 tcttctttta atatactttt ttgtttatct tatttctaat actttcccta atctctttct
     4141 ttcagggcaa taatgataca atgtatcatg cctctttgca ccattctaaa gaataacagt
     4201 gataatttct gggttaaggc aatagcaata tttctgcata taaatatttc tgcatataaa
     4261 ttgtaactga tgtaagaggt ttcatattgc taatagcagc tacaatccag ctaccattct
     4321 gcttttattt tatggttggg ataaggctgg attattctga gtccaagcta ggcccttttg
     4381 ctaatcatgt tcatacctct tatcttcctc ccacagctcc tgggcaacgt gctggtctgt
     4441 gtgctggccc atcactttgg caaagaattc accccaccag tgcaggctgc ctatcagaaa
     4501 gtggtggctg gtgtggctaa tgccctggcc cacaagtatc actaagctcg ctctagacta
     4561 gacgattaaa ttacattata gggcccgttt aaacccgctg atcagcctcg actgtgcctt
     4621 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
     4681 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
     4741 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
     4801 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct
     4861 ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     4921 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     4981 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     5041 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     5101 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     5161 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     5221 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     5281 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg
     5341 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     5401 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     5461 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     5521 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     5581 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     5641 aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcagcacgt
     5701 gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa ggtgaggaac
     5761 taaaccatgg ccaagttgac cagtgccgtt ccggtgctca ccgcgcgcga cgtcgccgga
     5821 gcggtcgagt tctggaccga ccggctcggg ttctcccggg acttcgtgga ggacgacttc
     5881 gccggtgtgg tccgggacga cgtgaccctg ttcatcagcg cggtccagga ccaggtggtg
     5941 ccggacaaca ccctggcctg ggtgtgggtg cgcggcctgg acgagctgta cgccgagtgg
     6001 tcggaggtcg tgtccacgaa cttccgggac gcctccgggc cggccatgac cgagatcggc
     6061 gagcagccgt gggggcggga gttcgccctg cgcgacccgg ccggcaactg cgtgcacttc
     6121 gtggccgagg agcaggactg acacgtgcta cgagatttcg attccaccgc cgccttctat
     6181 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     6241 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac
     6301 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     6361 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc
     6421 tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     6481 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     6541 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     6601 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     6661 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     6721 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     6781 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     6841 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     6901 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     6961 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     7021 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     7081 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     7141 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     7201 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     7261 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     7321 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     7381 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     7441 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     7501 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     7561 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     7621 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     7681 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     7741 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     7801 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     7861 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     7921 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     7981 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     8041 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     8101 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     8161 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     8221 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     8281 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     8341 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     8401 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     8461 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     8521 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     8581 aaagtg
//
