Comment: early pupae
TCAGATCCCGCTAAGATGTACAACGCGAGATCGCGTGGCAGCGCTGCCAACGAGGGGATTTCCCCCAAAAGAATAGTCAC CATCGGCTCTCTGCTCCTAGTCCTTGGTTTAAACCAGGTGGCGTCCATGTCGCTGGATCGCCTGGCTCTCGAGAGGAACG AGCTGCCAGCAGATCAACAGCAGCAACAACAGCAGGATGTTCTTGTCGACAATATGAAGCTCACCTCCATACCATCCGCC GACTCATCGGACATGTATATGCTGGTGCCCGATACGGTGGCCAGTCAACTGGAAGACACCGAGCATGTCAATCGCAACTC CATCGAGGCCGATCCCGAATCGGACCCCTCCGGCTCATCCTCCTCCGACATGCTGATGCTGGAGAGTGATTCCAGCCCGG CTATGCAACTCGTAGAACTGCCCAGCGACATTACGGTAGCCGAATCTCAGCCAGAAACGGAGCACGACACGGAGAACGAT CAGCAGCAGCCCGAGTCTATCGAGGAACATGTGGTGCTGATGAAACCAGCTAGCCAGGAGGCACAGTTGGAGCAGACGGT GGACTTGGCCACCGAAGCGGCACCAGTTCCATTAAACGACGAATTGCCAGAGGATGAGGAATCTCCTGCTGCCACCGAGA GTGCCGTAGAGGAATTGGAAAAGGAATCCGAAGCGGCCATGGATGATCAAGTGCCCGAGGAATCGGAAATTCAGCCGGAA CAGGTGCAACCAGGGGAATATCAATCAGAATCTGATGGTGAACAGGCGGAAACGAAGCCAGAGATTGAAGCTCAGCCAGA AGTGGAAGCTCAGCCAGAAGCGGAAGCTCAGCCAGAAGCGGAGCCCCAACTAGAAGTGGAGCCACAACCAGAAGTTGAGT CCCAGCCTGAAGTGGAATCCCAGCCAGAAGTAGAAGCCCAACCTGAAGTGGAACCTCAGTCAGAAGTGGAATCCCAGCCT GAGGCTGAATCTCATTCAGAACCAGAAACACAGGCAGAAGTGGAAGCCCAGCCTGAAGTGGAATCCCTACCAGAGGCTGA ATCCCAGCCAGAAGCAGAGTCCCAGCCAGAAAGGGAACCCGAAGTTGAGGCTGAGAAAATCAGTGATAACGAGGTGGACA CCACGGAGGCATCGCTGATGGAAACCCTGGTCGAGGGCATTGAAGATGGTCTAACTGCCGCCATGGACAACCTGGTGCCC GAAGAATTGGCAGAAGCCAGTGATAAGCAGGAAACGGAGCTGGAAAGCGAAGATCAGCAGTCCCCTGTCACAGAAGCCAT AGAAGAGCAGGCAGTTCCCGAAATTGAGCAGGAAAAGGAAAAGGAACCCGAGCAGATAACCTTGGCCGATGAAACCGAGC AGGATAGTGCCCAGCCTTCAAATGAAGAGCCAGTCGAGATTGCCCCAGAGCAACACACTGAAGCGGAGATTGCTCCAGCT AAAATTCCAGAGGAAGGCAAGCCTGAGGAGGAAGTCCAAGTGGAGGAGGAGTCTAAGCCAGTTGAAGAATCTAAGCCCGA GGAGGAAATCAAGCAACAGGAAGATGCCAAGCCAGAGGAGGATGACCAGAATTATGCTGAGACTCAACCAGCTGTCACCG AAGTGAAACCAGAGGAAACTCCTGCTGATATTCCTGTCGAGATCCCTGCTGAGGTTCCTGCCGAAATTCCTGCTGAGATC CCTGCAGAAGTTCCTGCTGAAATCCCTGCCGAGAGTCCTGCTGAGATCGCTGCCGAAGTTCCTGCCGAGATCCCTGCTGA AATCCCTGCTGAGATCCCTGCTGAAACTCCTGCTGAAACCCATGCTGAAATCCCTGCTGATGTTCCAGCTCAAGTTGTAG CTGAAGCTCCAGCGGAAACCCCAGCTGAAACCCCAGCAGAAGTTCCTGCTGAAATCCCATCTAAAGTCCAAGATGAAATT CAATCCGACTCCACCCAAGCCGAACCCCAAGTAGAGAAGGAAGCCCAGCAGCCGGAGAAGGAAACCAAGCCCCTGGAATC GTCCCTGCTGACCACCATCATTCCCATGGTGATGCCCCAACCCCAGGTGCCATCGCAACCGCTTCAGGTCCAGGATAGTG TGCTAAACTATGTCAACGCCTCGTTCATGCAGCAGCAGCCATCGGAAACGGATCTGCCCAAGACGGAGGAGGAGTCACCA TTCAATGCTCATCTGCGCCGCTATGATGGCGCCCAAGTGTGGCGCATCGTGGTCCAGACCGACAAGGATAAGCGAATGGC CGACGAACTGCAGTCCAAATATGATGGCCAACTGTGGAAAGAGGTCAAGCAGGAGGTGGACTATCTGCTGAAGCCACAAG TTTTGGCCGCCGCCGAACGTCACATTCGCGCCGCTAATCTTTCCCGGATCGTTCTGATCGACAATTTGCAGCGTGTCATT GAAAAGGAGAATCCGCCAGCGGAGAAGATTGCCGAGCTGCAGAACCGCAAGGGACATCGCCTCACCTGGCAGGCCTACCA TCGTCTGGAGGACATTCACGGCTTCATTGACTATATGGCTAAAACCTATCCGGATATTTGCTCCACTGAGATCATTGGCT ACTCGGTGGAGAAGCGACCACTTAAGATTCTCAAAATTTCCAATGGCAATGCCCGTAATCCTGGCATCTGGATCGATGGT GGCATGCACGCACGCGAATGGATCAGTCCGGCCACGGTCACCTACATTGCCAACCAGCTGGTCGAGGGTTGGGAGGATCT GCCCGAGCACATGCGCAGCATCAACTGGTACATCCATCCGGTGGCCAATCCCGATGGCTATGAGTATTCTCACACAACCG ATCGACTCTGGCGCAAGAATATGCGTGCCCATGGTCGCCAGTGCCCCGGTGTGGATCTGAACCGTAACTTTGGCTATAAG TGGGGCGGCAAGGGCACCTCGGCCAGTCCCTGCTCGCAGACATATCGCGGCAGCAAGGCCTTCTCCGAGCCGGAGACCTT CTACATCTCCAAGTTCATCAGCGGCTTTCCGCGCGAGACCTTCCAGGCGTACCTCTCGTTCCACAGCTATGGCCAGTACA TTCTGTATCCTTGGGGCTACGACTACCAGCTAACCCAGGACCGTGCCGATCTCGATCGTGTGGCACGACAGGCCGGAACG AGCATCACCAAGTCGACGGGCGTGAAGTATACGGTGGGATCCTCCGCCACAACTTTGTATCCCGCCGCCGGTGGCTCCGA TGATTGGGCCAAGGGCATTGCGGGCATCAAGTATGCCTACACCATTGAAATGGGCGACACCGGACGGTATGGCTTCGTCC TGCCCGCCCGATTCATCCAGTACAACGGCAAGGATGGTGTGACATTCGCCGATACCGTGGCCAGGGCCATTGCCCAGGGA CGCGGAAAATCGGTGGCCAGGAACAGCAGTGGCAGCCGGAGGCGTGGCCACTAGTCCTGACACTTAGCTCAATGCGATTT TTTGTAAATCAGCCTAATGATTTTTTTTTTTTACTACTTATATGAATACTATTTATTTATTTATTTATTCTACTTGTAAC TAGACAAATTGAACACACAAAAAAGAACACACAAGAAAAAAAAAAAAAAAAAAA
Cloned cDNAs prepared from polyadenylated RNA isolated from larvae and early pupae.
Larvae and early pupae at various stages of development were collected from an isogenic y; cn bw sp (iso-1) strain.
RNA was poly(A)+ selected once. Oligo(dT) primed with XhoI site at end of primer for first strand synthesis; EcoRI adapter on 5' ends of clones; size fractionated on Sephacryl S-500--approximately 1-6kb. cDNA directionally cloned into EcoRI/XhoI-digested pOT2 plasmid. ~500,000 colonies were plated out and a mass plasmid DNA prep was done using Qiagen Plasmids containing the cDNA inserts were size selected on Sephacryl S-500 column in order to exclude plasmids with no insert. Early elutions were collected and transformed into DH5 alpha strain (Gibco/BRL).
