LOCUS       sequence-209336-        9414 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In situ hybridization and RNAi studies in planarians.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9414)
  AUTHORS   Accorsi A, Williams MM, Ross EJ, Robb SMC, Elliott SA, Tu KC, 
            Sanchez Alvarado A
  TITLE     Hands-On Classroom Activities for Exploring Regeneration and Stem 
            Cell Biology with Planarians
  JOURNAL   The American Biology Teacher 79(3):208-223. Mar 2017.
REFERENCE   2  (bases 1 to 9414)
  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: "The 
            American Biology Teacher 79(3):208-223. Mar 2017."
COMMENT     Sequence Label: Ddo_slit-1
FEATURES             Location/Qualifiers
     source          1..9414
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        22..40
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        98..116
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3998..4014)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(4072..4090)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      4095..4142
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(4149..4165)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      4229..4315
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4407..4434
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             complement(join(5265..6056,6057..6125))
                     /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     5888..5907
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6126..6230)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6257..6712
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6344..6363)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6554..6575
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6853..6869
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        6876..6894
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     6927..6943
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7272..7288)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7273..7289)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(7317..7335)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(7356..7372)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7369..7391)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7380..7396
                     /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)."
     promoter        complement(join(7404..7410,7411..7428,7429..7434))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7449..7470
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(7758..8346)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7838..7857)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(8493..9302)
                     /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     9216..9235
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     9294..9313
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 caggaaacag ctatgaccat gtaatacgac tcactatagg ggatcctcaa aaaacccctc
       61 aagacccgtt tagaggcccc aaggggttat gtggaagtaa tacgactcac tatagggcga
      121 attgggtacc gggcccggcc aattctaccc gtttacagct tcgaggaaga gtaacgctac
      181 ggcgatagaa gcgtccatta tcgcatttga actgtatttt gtgcgtggta tattttgagg
      241 gttgacaaca aggagataaa ctgttgtctg tagatctttg ataaaagttc ctgttgctat
      301 ctgcttgcat tcgattatta gagcaagttc ctttacaaac tcggtattgg aactttttta
      361 ttgataaaca attgtcatga gtatctggat ctttgataaa agttcgagtt gttttgccat
      421 gacatttgga aactcttttt ttacattttc tgccaacaaa tccatttttg caaacgcatt
      481 tatattttcg atttttctca tttacttcgt gacaaacgcc attattggaa caaacgcccg
      541 gcttgctaca tattgtgctt ttgactggca aacttttggt tgacttaatc ggtacttcta
      601 tgtcttgtgt tggaacttct gacgatggta attcttgtgc ggttttcgat attaaatctc
      661 cacaaccggg tttaacgttc acggactttg acatgtattt ggaaaagtcg atttttttcc
      721 ggttgataag caacgattcg atgcatccag taaaatctga gtcgttatcg aaatgatggt
      781 tttgtattgc agtattaagg atttttttcg gtgctccacc aataaatatc ccggtgtcga
      841 gagccatata ttgattggtc attaggtact tggacttcag agtgaagtgt actttcccat
      901 tgaggtacat tgttaaattt tcaccaacta tttcgatatt cagataatgt aatttattat
      961 cattgaccac gtccattgaa aaaccatcgg gtccgatatc tttatcgcca atgttataac
     1021 ttatttttat atgtcctcga aatagattta caaccaagta gctcttatcg aacccttgat
     1081 agagaatcaa tccggatgaa gaattagtta taaattttag ggaaatgttt ccttttggca
     1141 ttaaaaatcc tcttgaaggt ggatgaattt caatatgtga gtctttaccg aaatataaac
     1201 tatttaaata ctgacagaaa ggacccgaat atcctggctt gcatttacaa tacgactttt
     1261 gtttattcat atcttcattt acacaaactc cgttattcaa acattcatga tatgcgcaaa
     1321 cagattcaga tattacttca tttatggatg tgaaatgctt tgagacttca cttctatcaa
     1381 tatctctcct ggaaagattc gtttttagtt tatacggttt ccatgctggt tcttcacagt
     1441 aagttccaac ataacccggt ctgcatatac atttgtaacc atttaccaag tcctcacatt
     1501 ttgcttcatt tttgcaaaca ttgtacttac aatcatcgat attttgttca cagttttttc
     1561 cttcccatcc ttttgtacaa gaacatttat agtcattatt ggaaagcggt tcacaatgtc
     1621 cagaattctc gcagggattt ttatcgtggc aatagaggaa tttgctctca caaaactttc
     1681 ctctgaaacc atttggacat ttacagctgt atttattcac ttggtctaca caggttgcac
     1741 cattttcaca tttgtgacct ccacaatcgt ctatattctc agtgcatttt ggaccacgga
     1801 aaccactggg acaaacacat tcataatttc cgaaattatc agtattttta caaatacctc
     1861 catttctaca cggttgacca aaacaagcat ttattctttt ttcacaactt ttcccatgaa
     1921 aaggattagg acaatcgcaa atgtaatcat gatgtgtaat cagtgaacat ttaccgttat
     1981 tttgacacgg agttttcata caagcgtcac atttggcagt gatataagtg tcttcggttt
     2041 cttctgaaca gataaactca gtgggttttg aatgataaat tgattttaat ttcatccttt
     2101 caggactggc acagagggaa atcccattgt ccaaaaatct gtccaaaaag aactgattca
     2161 gccattttat attacaatca caatggaatg ggttttgacc aagagcaatg ttattaagat
     2221 ttgttaagtc tcgaaatgct tgaaacggga tcgtagaaac atcattactt tgaagtatta
     2281 agacttttaa atttgaaagg tccttgaaag catcttgatg tatgcattgc aatttattga
     2341 aactcaaaac taatgtttcc aaatttttca aaggggtcaa cacatcttta ttgatttctg
     2401 tgataccatt ataactgagg accaaagttt ttagatttat tagattttta aatgatgatt
     2461 cattaagggt tttgagatca tttctgtcga gatatagttc cgtcgtattt ggaggaattt
     2521 ctgttgggat ttccttcagc tgaagttcac tacaatcgat tctcgtggat tcacatttgc
     2581 atttagacgg acactttgga attttattgc atttcatgtc taaacagcaa gaatagctaa
     2641 aattagcttt agcacatgaa gatgtctctt ccttcgaaga tttgttacct tcacatacaa
     2701 aatacttttc atccaaatta ggaagcggag ttcctttcaa aattggcggt gattggcaag
     2761 ttggaagtct tagacttcca agaataccgt agttattccg cgaccattcg gataaccatt
     2821 tcagacgaca gtcacaatga agttcgttat tgtttaacat tagatatttc agatttgtat
     2881 taaatttaaa agaatttgtt ccgatgcagc taatatgatt gtcgttgagt tgtaatattt
     2941 ctagttctat taaatgtttg aacgtttcat tattgacgca ttttaatttg tttgaactga
     3001 ggtttatttc tttcaatttg gtgcttttgc taaacgtgtc cggtttaata ttttcaattt
     3061 gattgttaga caagtctaat ttaactaaat tcgatgcctg atgaactgaa taaatttgat
     3121 ctaagttttt gatgtcattg tgagataaaa ttagagtggt cgttgaaggg aagatctttt
     3181 caggaagcga actcagaccg actcccgaac agttgacgat ttctgaagag caagagcatt
     3241 tatgtggaca aatgaccttc ggaacacagt tttcatcatg agtatcgtat ttaatatcag
     3301 attcagaatt acactgaaaa ttgtaattct ggagactctt tacggctatt ccatgagcgc
     3361 tactgggacc aacacatgat gccactgggg tcgcattgac ttccttgagt ttcaaaactt
     3421 gaactaacca cagtattttg cagtcacaat acaaaggatt gtggaccaga tatagatatc
     3481 ttagttttac gagaggtctc aggacttctt ccttcagagc agaaagttta ttttctgaca
     3541 aatgcaacaa gtaaagtctc ttgacccctt cgaatgtacc ggttggcagg cattcaattt
     3601 cgttattgtg taaatataaa aacattaatg aagacagtcc gttgaaagca tttcgtctta
     3661 tcgttcttaa attgttcatg tttaaatata aagccgttag ttgctgtaaa ccgataaatg
     3721 ctcctggttc aatgtcgacc attccatttt gcgttaaaat tatttttttc aataatttat
     3781 attttgatag aaatccttgt ctgagaatcg tgatgctatt attttgtaaa tctagttcaa
     3841 tggtatttgt cggaagatcc tgggggatgt tcgtaagctg tttaccagtg caattaacga
     3901 tcccgtcttg agaacatgtg cattgttttg gacacacttt tctgtctggt tgacaggtgt
     3961 tttcgggatg gtaatgccgc tagccatgga accggtggat ccactagttc tagagcggcc
     4021 gccaccgcgg tggagctcga attcatcgat gatatcagat ctgccggtct ccctatagtg
     4081 agtcgtatta cttcctagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     4141 tgcttaagac tggccgtcgt tttacaacgt cgtgactggg aaaacatcca tgctagcgtt
     4201 aacgcgagag tagggaactg ccaggcatca aataaaacga aaggctcagt cggaagactg
     4261 ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc ctgagtagga caaatccgcc
     4321 gggagcggat ttgaacgttg tgaagcaacg gcccggaggg tggcgggcag gacgcccgcc
     4381 ataaactgcc aggcatcaaa ctaagcagaa ggccatcctg acggatggcc tttttgcgtt
     4441 tctacaaact cttcctggct agcggtacgc gtattaattg cgttgcgctc actgcccgct
     4501 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     4561 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     4621 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     4681 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     4741 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaac
     4801 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     4861 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     4921 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     4981 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     5041 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     5101 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     5161 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     5221 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     5281 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     5341 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     5401 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     5461 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     5521 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     5581 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     5641 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     5701 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     5761 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     5821 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     5881 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     5941 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6001 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6061 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6121 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     6181 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     6241 cgaaaagtgc cacctgacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     6301 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     6361 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     6421 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     6481 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     6541 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     6601 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     6661 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat ttccattcgc
     6721 cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc
     6781 agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc
     6841 agtcacgacg ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat
     6901 tggagctcca ccgcggtggc ggccgctcta gaactagtgg atcccccgct ttgctgacac
     6961 ggtgtcactg cgctgaagag gctgtttgac gccatagtcc ctcctttttt ttccagatag
     7021 aacttgtgcc ctctgttgta aatactgctt gcagccttaa atattcctgc atggactgtg
     7081 gttagagcac tcactgactc ggttggtctc ggtggctggt agtgacctgc agagattaac
     7141 catttaaacg ccagagcctt cttgcagtct tcctggagaa agtttgcacc gcacttgtgg
     7201 ttctgtggtt gtttgtttta gtgagggtta ataagcggcc gcgggctgca ggaattcgat
     7261 atcaagctta tcgataccgt cgacctcgag ggggggcccg gtaccagctt ttgttccctt
     7321 tagtgagggt taatttcgag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
     7381 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
     7441 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
     7501 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     7561 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     7621 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     7681 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     7741 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     7801 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     7861 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     7921 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     7981 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     8041 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     8101 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     8161 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     8221 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     8281 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     8341 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgacgcgtaa
     8401 ctcacgttaa gggattttgg tcatgagctt gcgccgtccc gtcaagtcag cgtaatgctc
     8461 tgccagtgtt acaaccaatt aaccaattct gattagaaaa actcatcgag catcaaatga
     8521 aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt
     8581 aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct
     8641 gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg
     8701 ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg caaaagttta
     8761 tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc
     8821 gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcgatcg
     8881 ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc
     8941 gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa tgctgttttt
     9001 ccggggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg
     9061 gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc tgtaacatca
     9121 ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac
     9181 aagcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat
     9241 aaatcagcat ccatgttgga atttaatcgc ggcctcgacg tttcccgttg aatatggctc
     9301 ataacacccc ttgtattact gtttatgtaa gcagacagtt ttattgttca tgatgatata
     9361 tttttatctt gtgcaatgta acatcagaga ttttgagaca cgggccagag ctgc
//
