LOCUS       sequence-208380-        4305 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 4305)
  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 4305)
  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: Gsp_frz-4
FEATURES             Location/Qualifiers
     source          1..4305
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      complement(24..71)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        76..94
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     152..168
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     152..168
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(1935..1953)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(2011..2029)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2034..2050)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2034..2050)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2152..2171)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2163..2972
                     /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(2230..2249)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3119..3707
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3608..3627
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3861..3878
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(4037..4064)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(4156..4242)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(4291..4305,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(4305,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtctt aagcaaaaaa cccctcaaga cccgtttaga ggccccaagg
       61 ggttatgcta ggaagtaata cgactcacta tagggagacc ggcagatctg atatcatcga
      121 tgaattcgag ctccaccgcg gtggcggccg ctctagaact agtggatcca ccggttccat
      181 ggctagcggc attaccatcc cgagactccg agattacaat gctttgatca atatacgatg
      241 tagtaattat ttgaagcttt atctttgttc aatttatttt ccaatgtgtg agactgatac
      301 tagtatgaat tcacaaatcg tcaaaccatg caaacatctc tgtaaacatg taaaacaaaa
      361 atgcgagcca tttatgaaag aatctctaca aatgccttgg ccggaagaat tgaattgtga
      421 aaagttacca gaaaaaggaa aaatctgttt tacaccagaa tcgttttcac ttgatgattt
      481 agaccaacaa aatcagtctc aaggagatga tgttcaacaa ctttatcggc aatttgaaaa
      541 cgttatgcaa ggccataaaa ttaatcaatc aaaaattgaa atttttaaga aaatcggtgg
      601 aatgattggt tcacaatgtc taccatcaga aatcaatatt ggtcaattac cgaatatttc
      661 ttgtgctcag atctgttctg ctaatgtcca attcaatagt catgataaaa cattcgccaa
      721 tatttggttg atgggttggt cggttctttg ttttttttca tgttttctaa cgattattgt
      781 tttctcttgt aatacgaatc gatttctcta tccagaaaaa ccgattgttt tcctatcgat
      841 ttgttttttt ttctatgcta gtggtaattt attaggttca gtactcggtc gaaaagttgt
      901 tgcctgtcga tcatacgatg atcaaaatga ttttgttatc ggacaaggga gagagacaac
      961 atggtgcaaa atcaattttc taatggtcta ttttttcggt tcttctagtg ccttgtggtg
     1021 ggttgtccta actattactt ggtttctttc agcttctcgg cattggggct atgaagctat
     1081 cgaatcaatt agtagtatgt tgcatttact atgttgggca gttccagcgt taaaatcgat
     1141 atttatatta attctccata agattgatgc cgacgaatta accggtcaat gttatgtcgg
     1201 taataataat caaaatgttt tatggggttt tgtaattatt cctcaaatga tctatttgtt
     1261 gactggagtg atatttttga cattaggcta tatttcttta ttgaaagttc gaaaatcttt
     1321 tcttcaaaga ccagactgta tcgatacaaa taatttgaaa cgattagaaa aattaatggc
     1381 gaaaatcggt gttttttcaa tactttatat tgtaccggtg atttgtacta ttgctagtta
     1441 catggttgat gcatttaaaa tgacatattt cgatgttgct cttagagctt tatttagatt
     1501 ttctccaaat tgtgtcacga aaaatggggt taattggtcg agtataaagt gcattgaaat
     1561 tctacagccg atgctaccat cagttgaaat gagaatgttg agaatattta tgaatttagt
     1621 tattggtata acttccggaa tatggatatg gggaaactcg aaaacattga aatattgtat
     1681 aaaaaccatt agattttgga aacgaactaa agaactcgaa attcaaccag acaatcattg
     1741 cgactcatca acaactccaa acaattttcc agtagcccat gcacaatcaa aacaaaatga
     1801 aactctcttg ccttacaata actggaaatt gaattcttca tatccagcca atgaattcgg
     1861 tttagctact gtttcagtac ccagtgaaat tattcctccc gggtagaatt ggccgggccc
     1921 ggtacccaat tcgccctata gtgagtcgta ttacttccac ataacccctt ggggcctcta
     1981 aacgggtctt gaggggtttt ttgaggatcc cctatagtga gtcgtattac atggtcatag
     2041 ctgtttcctg gcagctctgg cccgtgtctc aaaatctctg atgttacatt gcacaagata
     2101 aaaatatatc atcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg
     2161 ttatgagcca tattcaacgg gaaacgtcga ggccgcgatt aaattccaac atggatgctg
     2221 atttatatgg gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc
     2281 gcttgtatgg gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg
     2341 ccaatgatgt tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc
     2401 cgaccatcaa gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc
     2461 ccggaaaaac agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg
     2521 atgcgctggc agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta
     2581 acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg
     2641 atgcgagtga ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa
     2701 tgcataaact tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg
     2761 ataaccttat ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa
     2821 tcgcagaccg ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt
     2881 cattacagaa acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc
     2941 agtttcattt gatgctcgat gagtttttct aatcagaatt ggttaattgg ttgtaacact
     3001 ggcagagcat tacgctgact tgacgggacg gcgcaagctc atgaccaaaa tcccttaacg
     3061 tgagttacgc gtcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     3121 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     3181 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     3241 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     3301 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     3361 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     3421 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     3481 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     3541 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     3601 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     3661 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     3721 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     3781 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
     3841 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac
     3901 gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc
     3961 ccgactggaa agcgggcagt gagcgcaacg caattaatac gcgtaccgct agccaggaag
     4021 agtttgtaga aacgcaaaaa ggccatccgt caggatggcc ttctgcttag tttgatgcct
     4081 ggcagtttat ggcgggcgtc ctgcccgcca ccctccgggc cgttgcttca caacgttcaa
     4141 atccgctccc ggcggatttg tcctactcag gagagcgttc accgacaaac aacagataaa
     4201 acgaaaggcc cagtcttccg actgagcctt tcgttttatt tgatgcctgg cagttcccta
     4261 ctctcgcgtt aacgctagca tggatgtttt cccagtcacg acgtt
//
