LOCUS       sequence-216997-        4062 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  An entry vector with U6a and U6c promoter driving rac2 guide RNAs 
            expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4062)
  AUTHORS   Zhou W, Cao L, Jeffries J, Zhu X, Staiger CJ, Deng Q
  TITLE     Neutrophil-specific knockout demonstrates a role for mitochondria in
            regulating neutrophil motility in zebrafish.
  JOURNAL   Dis Model Mech. 2018 Mar 28;11(3). pii: dmm.033027. doi: 
            10.1242/dmm.033027.
  PUBMED    29590639
REFERENCE   2  (bases 1 to 4062)
  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: "Dis Model 
            Mech."; date: "2018-03-28"; volume: "11(3). pii: dmm.033027. doi"; 
            pages: " 10.1242/dmm.033027"
COMMENT     Sequence Label: p3E-U6a-U6c-rac2 guide
FEATURES             Location/Qualifiers
     source          1..4062
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(624..747)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(761..777)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    793..927
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(817..836)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     871..890
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    complement(2171..2266)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2303..2322)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2304..2322)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2327..2343)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2327..2343)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2445..2464)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2456..3265
                     /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(2523..2542)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3412..4000
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3901..3920
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggccctg cagctctaga gctcgaattc tacaggtcac
      601 taataccatc taagtagttg attcatagtg actgcatatg ttgtgtttta cagtattatg
      661 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt
      721 ctcgttcaac tttcttgtac aaagttgaga tctataattc actggccgtc gttttacggt
      781 accatcgatg atgatccaga catgataaga tacattgatg agtttggaca aaccacaact
      841 agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta
      901 accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag
      961 gttcaggggg aggtgtggga ggttttttaa agcaagtaaa acctctacaa atgtggtatg
     1021 gctgattatg atcctctaga tcccttgggt agcagcgaag tcgaggcatt tctgtcctgg
     1081 ctggtctaga ggtctctcag gtttaaaaaa agcaccgact cggtgccact ttttcaagtt
     1141 gataacggac tagccttatt taaacttgct atgctgtttc cagcatagct cttaaacaca
     1201 gggtctcgca ttggcccgaa ctaggagcct ggagaactgc tatataaacc cgggacttct
     1261 gggtatgttt ctgccatgtg gtaagtggat aaacctactt attcaactca tttacctcaa
     1321 acatgacagt aaacctcttc ggggcaaacc tgcaaacgct taaaaccttc aaagtttgaa
     1381 accagcagtt ttgtagaggt ttaccgtagt atttttgaac tgtaaagcat atgcagagga
     1441 agctggtttt aggcttgtgt gattggatga gttcttctgt gacgtaactg atcttttgtc
     1501 ctcgtgtcct tcgccttccc ccacaggcat gcgcggacct tcccccgccg cagaccagag
     1561 gacaacatgc tcagacgcca gcaaacggcc catcgaaaca ttcacagact ctcggattgg
     1621 agagaccggg cccgaggtct ctgactaaaa aagcaccgac tcggtgccac tttttcaagt
     1681 tgataacgga ctagccttat ttaaacttgc tatgctgttt ccagcatagc tcttaaacgg
     1741 gactctggga tacagcccga accaagagct ggagggagag ctatatatac cagggacttc
     1801 tgggtatgtt tttgggaggt ggtgagtgac taaaccactt attcagctcc cttagatcaa
     1861 gtctgaccct atatcatggt gacataaacc tgcaaactga taaaacctga aggatctcaa
     1921 atccagagtt tgtgtgaggg attaccgtgg taattttcaa actgtaaagc atatgcaaaa
     1981 ttatctggtc ttggcttgag tgattgggtg tctcggtgtg atgcagggac gttttcagtg
     2041 acgtgtcctt ctccctcccc cacaggcatg cgcagaacat ttcccccctc cttgaagacc
     2101 agaacaaaag acgccgagag caggaaactc gtcttactga atgaccgagg ctggagaaat
     2161 cgaccctagg caactttatt atacaaagtt ggcattataa aaaagcattg cttatcaatt
     2221 tgttgcaacg aacaggtcac tatcagtcaa aataaaatca ttatttggag ctccatggta
     2281 gcgttaacgc ggccgcgata tcccctatag tgagtcgtat tacatggtca tagctgtttc
     2341 ctggcagctc tggcccgtgt ctcaaaatct ctgatgttac attgcacaag ataaaaatat
     2401 atcatcatga acaataaaac tgtctgctta cataaacagt aatacaaggg gtgttatgag
     2461 ccatattcaa cgggaaacgt cgaggccgcg attaaattcc aacatggatg ctgatttata
     2521 tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct atcgcttgta
     2581 tgggaagccc gatgcgccag agttgtttct gaaacatggc aaaggtagcg ttgccaatga
     2641 tgttacagat gagatggtca gactaaactg gctgacggaa tttatgcctc ttccgaccat
     2701 caagcatttt atccgtactc ctgatgatgc atggttactc accactgcga tccccggaaa
     2761 aacagcattc caggtattag aagaatatcc tgattcaggt gaaaatattg ttgatgcgct
     2821 ggcagtgttc ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt ttaacagcga
     2881 tcgcgtattt cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag
     2941 tgattttgat gacgagcgta atggctggcc tgttgaacaa gtctggaaag aaatgcataa
     3001 acttttgcca ttctcaccgg attcagtcgt cactcatggt gatttctcac ttgataacct
     3061 tatttttgac gaggggaaat taataggttg tattgatgtt ggacgagtcg gaatcgcaga
     3121 ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc cttcattaca
     3181 gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat tgcagtttca
     3241 tttgatgctc gatgagtttt tctaatcaga attggttaat tggttgtaac actggcagag
     3301 cattacgctg acttgacggg acggcgcaag ctcatgacca aaatccctta acgtgagtta
     3361 cgcgtcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     3421 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     3481 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     3541 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     3601 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     3661 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     3721 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     3781 actgagatac ctacagcgtg agcattgaga aagcgccacg cttcccgaag ggagaaaggc
     3841 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     3901 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     3961 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     4021 tttacggttc ctggcctttt gctggccttt tgctcacatg tt
//
