LOCUS       sequence-311727-        6272 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Tests for the impacts 4 uracils in a poly-uracil tract in 
            conjunctiuon with a hairpin 10 nts upstream have on human polymerase
            III transcription elongation from a U6 promoter. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6272)
  AUTHORS   Verosloff MS, Corcoran WK, Dolberg TB, Bushhouse DZ, Leonard JN, 
            Lucks JB
  TITLE     RNA Sequence and Structure Determinants of Pol III Transcriptional 
            Termination in Human Cells.
  JOURNAL   J Mol Biol. 2021 Jun 25;433(13):166978. doi: 
            10.1016/j.jmb.2021.166978. Epub 2021 Apr 1.
  PUBMED    33811918
REFERENCE   2  (bases 1 to 6272)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Mol 
            Biol."; date: "2021-06-25"; volume: "433(13):166978. doi"; pages: " 
            10.1016/j.jmb.2021.166978. Epub 2021 Apr 1"
COMMENT     Sequence Label: 
            pAV-U6+27-Linear-2_PolyU(4)_HairpinDistance(10)nts_Tornado-Corn
FEATURES             Location/Qualifiers
     source          1..6272
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     71..90
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     polyA_signal    597..718
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     repeat_region   767..896
                     /label=AAV2 ITR
     rep_origin      complement(990..1445)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1127..1148)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1339..1358
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1472..1576
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1577..1645,1646..2437)
                     /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(1795..1814)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2608..3196
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3097..3116
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3465..3599
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(4024..4044)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          4174..4239
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             complement(4659..5453)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIYCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4770..4789)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5380..5399
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(5810..6139)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5824..5959
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6124..6144
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(6172..6272,1..148)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6172..6192
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
ORIGIN
        1 acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt taaaattatg
       61 ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt tcttggcttt
      121 atatatcttg tggaaaggac gaaacaccgt gctcgcttcg gcagcacata tactaggttg
      181 cggctttctg ctgcaatcgt gattgatgtt ttcgacggcc atcagtcgcc ggtcccaagc
      241 ccggataaaa tgggaggggg cgggaaaccg cctaaccatg ccgactgatg gcaggcccgg
      301 atagctcagt cggtagagca gcggccgcgc gattcgagga aggaggtctg aggaggtcac
      361 tgaatcgcgc ggccgcgggt ccagggttca agtccctgtt cgggcctgcc atcagtcggc
      421 gtggactgta gaacactgcc aatgccggtc ccaagcccgg ataaaagtgg agggtacagt
      481 ccacgctcta gagcggactt cggtccgctt tttactagga cctgcaggca tgcaagcttg
      541 acgtcggtta ccgatatcca tatggcggcc gcatcgatct cgagccgcgg actagtaact
      601 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
      661 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
      721 atgtcttacg tagataagta gcatggcggg ttaatcatta actacaagga acccctagtg
      781 atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag
      841 gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagagag
      901 ggacagatct gcagatctgc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
      961 aacagttgcg cagcctgaat ggcgaatgga aattgtaagc gttaatattt tgttaaaatt
     1021 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     1081 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     1141 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     1201 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     1261 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     1321 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     1381 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     1441 cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     1501 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     1561 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     1621 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     1681 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     1741 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     1801 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     1861 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     1921 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     1981 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     2041 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     2101 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     2161 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     2221 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     2281 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     2341 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     2401 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     2461 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     2521 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     2581 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     2641 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     2701 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     2761 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     2821 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     2881 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     2941 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     3001 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     3061 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     3121 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     3181 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     3241 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     3301 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     3361 gagcgagcgc gcagagaggg agtggccaac tccatcacta ggggttcctg gaggggtgga
     3421 gtcgtgacgt gaattacgtc atagggttag ggaggtcctg gatcgatcca gacatgataa
     3481 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
     3541 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
     3601 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt
     3661 aaagcaagta aaacctctac aaatgtggta tggctgatta tgatctctag tcaaggcact
     3721 atacatcaaa tattccttat taaccccttt acaaattaaa aagctaaagg tacacaattt
     3781 ttgagcatag ttattaatag cagacactct atgcctgtgt ggagtaagaa aaaacagtat
     3841 gttatgatta taactgttat gcctacttat aaaggttaca gaatattttt ccataatttt
     3901 cttgtatagc agtgcagctt tttcctttgt ggtgtaaata gcaaagcaag caagagttct
     3961 attactaaac acagcatgac tcaaaaaact tagcaattct gaaggaaagt ccttggggtc
     4021 ttctaccttt ctcttctttt ttggaggagt agaatgttga gagtcagcag tagcctcatc
     4081 atcactagat ggcatttctt ctgagcaaaa caggttttcc tcattaaagg cattccacca
     4141 ctgctcccat tcatcagttc cataggttgg aatctaaaat acacaaacaa ttagaatcag
     4201 tagtttaaca cattatacac ttaaaaattt tatatttacc ttagagcttt aaatctctgt
     4261 aggtagtttg tccaattatg tcacaccaca gaagtaaggt tccttcacaa agatccggac
     4321 caaagcggcc atcgtgcctc cccactcctg cagttcgggg gcatggatgc gcggatagcc
     4381 gctgctggtt tcctggatgc cgacggattt gcactgccgg tagaactccg cgaggtcgtc
     4441 cagcctcagg cagcagctga accaactcgc gaggggatcg agcccggggt gggcgaagaa
     4501 ctccagcatg agatccccgc gctggaggat catccagccg gcgtcccgga aaacgattcc
     4561 gaagcccaac ctttcataga aggcggcggt ggaatcgaaa tctcgtgatg gcaggttggg
     4621 cgtcgcttgg tcggtcattt cgaaccccag agtcccgctc agaagaactc gtcaagaagg
     4681 cgatagaagg cgatgcgctg cgaatcggga gcggcgatac cgtaaagcac gaggaagcgg
     4741 tcagcccatt cgccgccaag ctcttcagca atatcacggg tagccaacgc tatgtcctga
     4801 tagcggtccg ccacacccag ccggccacag tagatgaatc cagaaaagcg gccattttcc
     4861 accatgatat tcggcaagca ggcatcgcca tgggtcacga cgagatcctc gccgtcgggc
     4921 atgcgcgcct tgagcctggc gaacagttcg gctggcgcga gcccctgatg ctcttcgtcc
     4981 agatcatcct gatcgacaag accggcttcc atccgagtac gtgctcgctc gatgcgatgt
     5041 ttcgcttggt ggtcgaatgg gcaggtagcc ggatcaagcg tatgcagccg ccgcattgca
     5101 tcagccatga tggatacttt ctcggcagga gcaaggtgag atgacaggag atcctgcccc
     5161 ggcacttcgc ccaatagcag ccagtccctt cccgcttcag tgacaacgtc gagcacagct
     5221 gcgcaaggaa cgcccgtcgt ggccagccac gatagccgcg ctgcctcgtc ctgcagttca
     5281 ttcagggcac cggacaggtc ggtcttgaca aaaagaaccg ggcgcccctg cgctgacagc
     5341 cggaacacgg cggcatcaga gcagccgatt gtctgttgtg cccagtcata gccgaatagc
     5401 ctctccaccc aagcggccgg agaacctgcg tgcaatccat cttgttcaat catgcgaaac
     5461 gatcctcatc ctgtctcttg atcagatctt gatcccctgc gccatcagat ccttggcggc
     5521 aagaaagcca tccagtttac tttgcagggc ttcccaacct taccagaggg cgccccagct
     5581 ggcaattccg gttcgcttgc tgtccataaa accgcccagt ctagctatcg ccatgtaagc
     5641 ccactgcaag ctacctgctt tctctttgcg cttgcgtttt cccttgtcca gatagcccag
     5701 tagctgacat tcatccgggg tcagcaccgt ttctgcggac tggctttcta cgtgttccgc
     5761 ttcctttagc agcccttgcg ccctgagtgc ttgcggcagc gtgaagcttt ttgcaaaagc
     5821 ctaggcctcc aaaaaagcct cctcactact tctggaatag ctcagaggcc gaggcggcct
     5881 cggcctctgc ataaataaaa aaaattagtc agccatgggg cggagaatgg gcggaactgg
     5941 gcggagttag gggcgggatg ggcggagtta ggggcgggac tatggttgct gactaattga
     6001 gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca cacctggttg
     6061 ctgactaatt gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc
     6121 cacaccctaa ctgacacaca ttccacagcc ggatccaagg tcgggcagga agagggccta
     6181 tttcccatga ttccttcata tttgcatata cgatacaagg ctgttagaga gataattaga
     6241 attaatttga ctgtaaacac aaagatatta gt
//
