LOCUS       sequence-374766-        3148 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CR3 constant region - hU6 sgRNA promoter flanked by BsmBI sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3148)
  AUTHORS   Replogle JM, Bonnar JL, Pogson AN, Liem CR, Maier NK, Ding Y, 
            Russell BJ, Wang X, Leng K, Guna A, Norman TM, Pak RA, Ramos DM, 
            Ward ME, Gilbert LA, Kampmann M, Weissman JS, Jost M
  TITLE     Maximizing CRISPRi efficacy and accessibility with dual-sgRNA 
            libraries and optimal effectors.
  JOURNAL   eLife. 2022 Dec 28;11:e81856. doi: 10.7554/eLife.81856.
  PUBMED    36576240
REFERENCE   2  (bases 1 to 3148)
  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: "eLife."; 
            date: "2022-12-28"; volume: "11:e81856. doi"; pages: " 
            10.7554/eLife.81856"
COMMENT     Sequence Label: pJR98
FEATURES             Location/Qualifiers
     source          1..3148
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(114..133)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     233..255
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(293..311)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        379..483
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(484..552,553..1344)
                     /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(702..721)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1515..2103
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2004..2023
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2257..2274
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2391..2412
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2427..2432,2433..2450,2451..2457)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2465..2481
                     /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)."
     primer_bind     2470..2492
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2489..2505
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2489..2505
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2615..2634)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(2785..2805)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(3037..3054)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3037..3053)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3046..3068)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat
       61 tttctcctta cgcatctgtg cggtatttca caccgcattt ggtgcactct cagtacaatc
      121 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc
      181 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
      241 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg
      301 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
      361 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
      421 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggtag
      481 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
      541 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
      601 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
      661 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
      721 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
      781 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
      841 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
      901 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
      961 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     1021 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     1081 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     1141 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     1201 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     1261 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     1321 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     1381 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     1441 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     1501 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     1561 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     1621 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     1681 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     1741 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     1801 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     1861 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     1921 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcgacagggt cggaacagga
     1981 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     2041 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     2101 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     2161 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     2221 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     2281 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     2341 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt
     2401 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt
     2461 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccacg
     2521 agtcgcagca cagctcgctg gtccagaact gatcgtctcc tttctggctt tccacaagat
     2581 atataaagcc aagaaatcga aatactttca agttacggta agcatatgat agtccatttt
     2641 aaaacataat tttaaaactg caaactaccc aagaaattat tactttctac gtcacgtatt
     2701 ttgtactaat atctttgtgt ttacagtcaa attaattcca attatctctc taacagcctt
     2761 gtatcgtata tgcaaatatg aaggaatcat gggaaatagg ccctcttcct gcccgacctt
     2821 ggggcgcgcc atagctgagt gtagattcga gcaaaaaaag cacccgactc gggtgccact
     2881 ttttcaagtt gtaaacggac tagccttatt tcaacttgct atgcactctt gtgcttagct
     2941 cttgagacga tctgtcgccc gtctcaaacg caaccctccg gcggcggcga tcgggtcagc
     3001 tcatcgatga atcgctatcc atcgtcatcg cgttgcactg gccgtcgttt tacaacgtcg
     3061 tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc
     3121 cagctggcgt aatagcgatg aggcccgc
//
