LOCUS       sequence-274588-        5654 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Human ILVBL poly(A) site reporter, stem loop mutant rescue.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5654)
  AUTHORS   Wu X, Bartel DP
  TITLE     Widespread Influence of 3'-End Structures on Mammalian mRNA 
            Processing and Stability.
  JOURNAL   Cell. 2017 May 18;169(5):905-917.e11. doi: 
            10.1016/j.cell.2017.04.036.
  PUBMED    28525757
REFERENCE   2  (bases 1 to 5654)
  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"; doi: 
            "10.1016/j.cell.2017.04"; journalName: "Cell"; date: "2017-05-18- 
            18"; volume: "169"; issue: "5"; pages: "905-917.e11"
COMMENT     Sequence Label: pCMV-GFP-ILVBLpA-G10C-C21G
FEATURES             Location/Qualifiers
     source          1..5654
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        347..550
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     501..521
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    573..653
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      655..664
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(661..663,664..666,667..1380)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(661..663,664..666,667..1377)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(706..727)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(967..986)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1314..1335
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2642..2661)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2707..2762
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2761..2780)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3123..3139)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3123..3139)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3136..3158)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3147..3163
                     /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(3171..3177,3178..3195,3196..3201))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3216..3237
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3296..3491
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3342..3477
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3497..3631
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3534..3553)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3588..3607
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3699..3716)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3870..4458)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3950..3969)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4629..5420,5421..5489))
                     /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     5252..5271
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5490..5594)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        join(5625..5654,1..346)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
       61 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
      121 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
      181 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      241 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
      301 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
      361 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
      421 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
      481 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
      541 aagcagagct ggtttagtga accgtcagat ccgctagcgc taccggactc agatctcgag
      601 ctcaagcttc gaattctgca gtcgacggta ccgcgggccc gggatccacc ggtcgccacc
      661 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
      721 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
      781 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
      841 ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag
      901 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
      961 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     1021 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     1081 aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac
     1141 ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
     1201 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     1261 tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     1321 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa
     1381 agcggccgca ctcctcaggt gcgatcaggt ggtcaaggtg ctgcacgatg cccagcagca
     1441 gtgccgagac ggccacccgg ttgtggtcaa catcctcatt gggaggacgg acttccgcga
     1501 tggctccatt gctgtatagg gccttgtggg tcaggacgct tggctgcctt cctgcccctg
     1561 gactgtgtcc agctggtttg gagtctcatc attgcttgcc ctgggcctat cccatgaggc
     1621 ccagtgactc cacaacctcc ctgaggaggg gatggagggg tcaaatctca gagaggctct
     1681 cttgtgagct cttggaccct cctcttcacg gacccaactg tgccgtttgt cccctctctt
     1741 atggagactg aataaaccac ctctgcccca tgtagggcca agtactcagt ctcagaacct
     1801 gtgcatattt atttattatc cacaaagatg gaggtgcgga aaagaagagg gagacaggag
     1861 ggaaccgtac tctcccactg gaactctggg gcccactgag gcaccattat tggggatttc
     1921 agggtggctg ggcactgcaa actgctccct cctctgtggt ccctgaaaga gccccacacg
     1981 cctgcttcag acgtgtccac gcacaccagt cctcacagac acacacacac atgcatggag
     2041 gcaataaata tgttccgtac cagactgccc ccagcctgac gcttcagggg gccccctcca
     2101 aaagggaggg gtttaagtgc tcaatttttt tcgggggggg ggcaaggggg gggcaaggag
     2161 atggggattg gaaaggccag actctgttat ctccatttgc tgactagagg ccaatcgtgg
     2221 ggtcttcccc ggaaaagggt aggggaaacc agctgtttct ctataagccc caggctaccc
     2281 ctgtgtccat gctggaaccc aggatagggg ctgcccccca caaacagtgc ctcatgctag
     2341 tctttgtgtc gtccaaggga gggagaaaca gtgcagtgct caaagctaat actgatttgg
     2401 ttggcagcag caagagggcc gactagaggt gactcaccac accagggtga tgctcacagt
     2461 aactggtgtt gccccggcca aggggaggca agagaggcca gcctagcaag cgcgagaact
     2521 acctttttgg caacaaaagg ggaagccctt ggtgctcagg aaagatggtc acaaccgagg
     2581 ctgcctatca gaaggtggtg gctggtgtgg ccaatgccct ggctcacaaa taccactgag
     2641 atctttttcc ctctgccaaa aattatgggg acatcatgaa gccccttgag catctgactt
     2701 ctggctaata aaggaaattt attttcattg caatagtgtg ttggaatttt ttgtgtctct
     2761 cactcggaag gacatatggg agggcaaatc atttaaaaca tcagaatgag tatttggttt
     2821 agagtttggc aacatatgcc catatgctgg ctgccatgaa caaaggttgg ctataaagag
     2881 gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga aaagccttga
     2941 cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta acatccctaa
     3001 aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta ctcccagtca
     3061 tagctgtccc tcttctctta tggagatccc tcgacctgca gcccaagctt ggcgtaatca
     3121 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     3181 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3241 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagcg gatccgcatc
     3301 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     3361 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     3421 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3481 gcttttgcaa aaagctaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     3541 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     3601 actcatcaat gtatcttatc atgtctggat ccgctgcatt aatgaatcgg ccaacgcgcg
     3661 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     3721 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     3781 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     3841 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     3901 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     3961 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4021 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4081 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4141 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4201 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4261 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     4321 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4381 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     4441 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4501 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     4561 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     4621 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     4681 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     4741 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     4801 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     4861 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     4921 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     4981 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     5041 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5101 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5161 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5221 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5281 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5341 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     5401 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     5461 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     5521 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     5581 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg ggtcgacatt gattattgac
     5641 tagtaatagt aatc
//
