LOCUS       sequence-404040-        7526 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Backbone for cloning splicing library with truncated CAG promoter 
            and first half of beta globin minigene reporter (exon 1 and intron 
            1)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7526)
  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 7526)
  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_Backbone
FEATURES             Location/Qualifiers
     source          1..7526
                     /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(3546..3563)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3552..3776
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3822..4250
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3909..3928)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4119..4140
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4259..4279)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4264..4593
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4444..4579
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4506..4525
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4641..4688
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4707..5081
                     /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    5211..5332
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5248..5267)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5302..5321
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5381..5397)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5381..5397)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5394..5416)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5405..5421
                     /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(5429..5435,5436..5453,5454..5459))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5474..5495
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5612..5629)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5783..6371)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5863..5882)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6542..7333,7334..7402))
                     /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     7165..7184
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7403..7507)
                     /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 ttctctgaga cgaatctaga gggcccgttt aaacccgctg
     3541 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     3601 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     3661 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     3721 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     3781 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     3841 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     3901 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     3961 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     4021 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     4081 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     4141 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     4201 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     4261 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     4321 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     4381 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     4441 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     4501 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     4561 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     4621 ttcggatctg atcagcacgt gttgacaatt aatcatcggc atagtatatc ggcatagtat
     4681 aatacgacaa ggtgaggaac taaaccatgg ccaagttgac cagtgccgtt ccggtgctca
     4741 ccgcgcgcga cgtcgccgga gcggtcgagt tctggaccga ccggctcggg ttctcccggg
     4801 acttcgtgga ggacgacttc gccggtgtgg tccgggacga cgtgaccctg ttcatcagcg
     4861 cggtccagga ccaggtggtg ccggacaaca ccctggcctg ggtgtgggtg cgcggcctgg
     4921 acgagctgta cgccgagtgg tcggaggtcg tgtccacgaa cttccgggac gcctccgggc
     4981 cggccatgac cgagatcggc gagcagccgt gggggcggga gttcgccctg cgcgacccgg
     5041 ccggcaactg cgtgcacttc gtggccgagg agcaggactg acacgtgcta cgagatttcg
     5101 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     5161 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     5221 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     5281 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     5341 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     5401 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     5461 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     5521 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     5581 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     5641 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     5701 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     5761 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     5821 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     5881 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     5941 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     6001 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     6061 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     6121 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     6181 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     6241 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     6301 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     6361 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     6421 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     6481 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     6541 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     6601 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     6661 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     6721 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     6781 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     6841 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     6901 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     6961 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     7021 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     7081 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     7141 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     7201 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     7261 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     7321 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     7381 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     7441 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     7501 ttccgcgcac atttccccga aaagtg
//
