LOCUS       sequence-215035-        2849 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PCR template vector for amplifying gRNAcore-pU6.2 promoter fragment.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2849)
  AUTHORS   Poe AR, Wang B, Sapar ML, Ji H, Li K, Onabajo T, Fazliyeva R, Gibbs 
            M, Qiu Y, Hu Y, Han C
  TITLE     Robust CRISPR/Cas9-Mediated Tissue Specific Mutagenesis Reveals Gene
            Redundancy and Perdurance in Drosophila.
  JOURNAL   Genetics. 2018 Nov 30. pii: genetics.118.301736. doi: 
            10.1534/genetics.118.301736.
  PUBMED    30504366
REFERENCE   2  (bases 1 to 2849)
  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: "Genetics. 
            2018 Nov 30. pii: genetics.118.301736. doi: 
            10.1534/genetics.118.301736."
COMMENT     Sequence Label: pTGC-U6.3
FEATURES             Location/Qualifiers
     source          1..2849
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(56..142)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     191..213
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     205..222
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     206..222
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_RNA        245..320
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        322..742
                     /label=dU6-3 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-3 
                     snRNA (Port et al., 2014)"
     primer_bind     complement(759..778)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(760..778)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(783..799)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(783..799)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(901..920)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             912..1721
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(979..998)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      1868..2456
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2357..2376
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2610..2627
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(2786..2813)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ctgcccgcca ccctccgggc cgttgcttca caacgttcaa atccgctccc ggcggatttg
       61 tcctactcag gagagcgttc accgacaaac aacagataaa acgaaaggcc cagtcttccg
      121 actgagcctt tcgttttatt tgatgcctgg cagttcccta ctctcgcgtt aacgctagca
      181 tggatgtttt cccagtcacg acgttgtaaa acgacggcca gtcttaagct cgggccgctc
      241 ttctgtttta gagctagaaa tagcaagtta aaataaggct agtccgttat caacttgaaa
      301 aagtggcacc gagtcggtgc tttttttgct cacctgtgat tgctcctact caaatacaaa
      361 aacatcaaat tttctgtcaa taaagcatat ttatttatat ttattttaca ggaaagaatt
      421 ccttttaaag tgtattttaa cctataatga aaaacgatta aaaaaaatac ataaaataat
      481 tcgaaaattt ttgaatagcc caggttgata aaaattcatt tcatacgttt tataacttat
      541 gcccctaagt attttttgac catagtgttt caattctaca ttaattttac agagtagaat
      601 gaaacgccac ctactcagcc aagaggcgaa aaggttagct cgccaagcag agagggcgcc
      661 agtgctcact actttttata attctcaact tctttttcca gactcagttc gtatatatag
      721 acctattttc aatttaacgt cgggaagagc ctgatatccc ctatagtgag tcgtattaca
      781 tggtcatagc tgtttcctgg cagctctggc ccgtgtctca aaatctctga tgttacattg
      841 cacaagataa aaatatatca tcatgaacaa taaaactgtc tgcttacata aacagtaata
      901 caaggggtgt tatgagccat attcaacggg aaacgtcgag gccgcgatta aattccaaca
      961 tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga
     1021 caatctatcg cttgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag
     1081 gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta
     1141 tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg ttactcacca
     1201 ctgcgatccc cggaaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa
     1261 atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttgtaatt
     1321 gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga atgaataacg
     1381 gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct
     1441 ggaaagaaat gcataaactt ttgccattct caccggattc agtcgtcact catggtgatt
     1501 tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac
     1561 gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt
     1621 tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga
     1681 ataaattgca gtttcatttg atgctcgatg agtttttcta atcagaattg gttaattggt
     1741 tgtaacactg gcagagcatt acgctgactt gacgggacgg cgcaagctca tgaccaaaat
     1801 cccttaacgt gagttacgcg tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     1861 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     1921 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     1981 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     2041 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     2101 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     2161 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     2221 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     2281 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     2341 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     2401 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     2461 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     2521 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     2581 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     2641 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     2701 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatacg cgtaccgcta
     2761 gccaggaaga gtttgtagaa acgcaaaaag gccatccgtc aggatggcct tctgcttagt
     2821 ttgatgcctg gcagtttatg gcgggcgtc
//
