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===Notes and primers used etc=== Blocking primer for protecting 5' end from RNase III degradation T7-frag-block 5'- CTGTCTCTTATACACATCTTAATGTCTCCC/3InvdT/ 3InvdT so don't get any nonspecific extension by MMLV RT. Stuff used for RNA-seq for BENG160 class 5β end addition primers TSO_N10_BC[XX] [AAGCAGTGGTATCAACGCAGAGdUdU]NNNNNNNNNNTTTAGGrGrGrG adatpor1 5'- AAG CAG TGG TAT CAA CGC AGA G/ideoxyU//ideoxyU/ NNN NNN NNN NTT TAG GrGrGrG -3' P1-STRT 5β- [AATGATACGGCGACCACCGA][GATCT][AAGCAGTGGTATCAACGCAGAGT] -3β (Tm=82.59) ILA adaptor blue Tm=64.31 adaptor1 Tm=61.02 STRT-SEQ 5'- [GATCT][AAGCAGTGGTATCAACGCAGAGTT] -3' adaptor1 3β end addition primers T20VN_PE_R 5'-Bio-[GCATTCCTGCTGAACCGCTCTT]CCGATCTTTTTTTTTTTTTTTTTTTTTVN -3β (Tm=78.92) adaptor2 Tm=65.68 PCR_R.N2Ind[XX] (XX=23,24 for me) 5β- [CAAGCAGAAGACGGCATACGAGAT][TACAAG]CTCG][GCATTCCTGCTGAACCGCTCTT] -3β (Tm=87.12) ILA adaptor orange bc adaptor2 Tm=65.68 My modifications for T7-tspns for 5' addition primers To use with (T7-top and T7-top2) transposons 5β- [AATGATACGGCGACCACCGA][GATCT][CTCCCTCGCGCCATCAGAGAT] -3β (Tm=86.59) ILA adaptor blue Top2-5βend (T7-top2-PCR-iaf) Tm=66.79 (can be used if top1 is used for transposition) To use with (T7-top3) transposon 5β- [AATGATACGGCGACCACCGA][GATCT][GGGAGACATTAAGATGTGTATAAGAGACAG] -3β (Tm=81.40) ILA adaptor blue Top3-5βend (T7-top3-PCR-iaf) Tm=60.71
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