LOCUS       sequence-442846-        3481 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Plasmid to express gRNA2 (aatttcgacgggccttcaag) for editing at the 
            end of Drosophila 5-HT1BR coding frame.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3481)
  AUTHORS   Mallick A, Tan HL, Epstein JM, Gaudry Q, Dacks AM
  TITLE     Serotonin acts through multiple cellular targets during an olfactory
            critical period.
  JOURNAL   bioRxiv [Preprint]. 2024 Apr 14:2024.04.14.589413. doi: 
            10.1101/2024.04.14.589413.
  PUBMED    38645269
REFERENCE   2  (bases 1 to 3481)
  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: "bioRxiv 
            [Preprint]. 2024 Apr 14:2024.04.14.589413. doi: 
            10.1101/2024.04.14.589413."
COMMENT     Sequence Label: pdU6-2_sgRNA2-5-HT1BR
FEATURES             Location/Qualifiers
     source          1..3481
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     182..201
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     435..452
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    569..590
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(605..610,611..628,629..635)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    643..659
                     /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     648..670
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     667..683
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     667..683
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     702..722
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        704..722
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_RNA        complement(853..928)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(950..1349)
                     /label=dU6-2 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-2 
                     snRNA (Das et al., 1987)"
     primer_bind     complement(1369..1388)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1370..1388)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1398..1415)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1398..1414)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1407..1429)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(1555..2011)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1693..1714)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1905..1924
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2038..2142
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2143..2211,2212..3003)
                     /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(2361..2380)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(3174..3481,1..281)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
       61 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
      121 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
      181 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
      241 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
      301 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
      361 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
      421 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac
      481 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact
      541 ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc
      601 aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat
      661 ttcacacagg aaacagctat gaccatgatt acgccaagcg cgcaattaac cctcactaaa
      721 gggaacaaaa gctggagctc actagtaatt ctttcctgta aaataaatat aaataaatat
      781 gctttattga cagaaaattt gatgtttttg tatttgagta ggagcaatca caggtgagca
      841 aaaaaatgga tcgcaccgac tcggtgccac tttttcaagt tgataacgga ctagccttat
      901 tttaacttgc tatttctagc tctaaaacct tgaaggcccg tcgaaattcg aagtattgag
      961 gaaaacatac ctatataaat gatcaacatc aggaaagagc agttgagaat tataagaatt
     1021 ggcaaatggt cttaagaacc ctctgcttaa gattttcaaa atttccttta gaatcaaagt
     1081 gtcctattgt tcgtttgttg aaaacacagt tcgatttatt gactataata aattgatagt
     1141 tttaaatata gaggcacgac taagagagtt tggttttgtt ttggttaggg accaaaaaaa
     1201 gtatacataa cgaataaaaa aggatttaag tgcaaatggt aaaaaagtgt cgagtttttc
     1261 ttgagttgat tgtgctgtaa tgagactctg cattcggcat ttgactcggc ttttcctact
     1321 cgtgccgtat ttcaggctgc aagtcgaacc aagcttgtac ccaattcgcc ctatagtgag
     1381 tcgtattacg cgcgctcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc
     1441 gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa
     1501 gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggaaattg
     1561 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcattttttt
     1621 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     1681 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     1741 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     1801 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     1861 tttagagctt gacggggaaa gccggcgaac gtggtgagaa aggaagggaa gaaagcgaaa
     1921 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
     1981 gccgcgctta atgcgccgct acagggcgcg tcaggtggca cttttcgggg aaatgtgcgc
     2041 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     2101 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     2161 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     2221 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     2281 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     2341 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     2401 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     2461 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     2521 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     2581 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     2641 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     2701 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     2761 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     2821 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     2881 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     2941 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     3001 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     3061 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     3121 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     3181 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     3241 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     3301 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     3361 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     3421 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     3481 c
//
