Illumina HiSeq 2000 sequencing; Transcriptional profiling of pancreatic beta cells during maturation and with function-inactivating mutations
Transcriptional profiling of pancreatic beta cells during maturation and with function-inactivating mutations
MytDelpanc and MipeGFP islets were collected from mice postnatal day 1. The islets from 2-3 mice of the same genotype were pooled for each sample. Islets were dispersed using trypsin and FACS purified based on eGFP expression. Propidium Iodine was used to select live cells. MytDelpanc genotype: Myt1tm1.1Ggu/tm1.1Ggu;Myt1Ltm1.1Ggu/tm1.1Ggu;St18tm1.1Ggu/tm1.1Ggu; Tg(Pdx1-cre)89.1Dam/+; Tg(Ins1-EGFP)1Hara/J. Myt1tm1.1Ggu/tm1.1Ggu information: http://www.informatics.jax.org/allele/genoview/MGI:3765058?counter=1 and Tg(Pdx1-cre)89.1Dam/+ allele information: http://www.informatics.jax.org/allele/MGI:2684317. The Myt1Ltm1.1Ggu allele has two loxP sites flanking the exons #12 and #13. Cre-mediated inactivation produces an allele that makes no Myt1L protein. The St18tm1.1Ggu allele has two loxP sites flanking the exons #9 and #10. Cre-mediated inactivation produces an allele that makes no St18 protein. Mice used in this experiment contained pancreatic cell specific inactivation of a family of genes, Myt1, Myt1L, and St18. These mutants carried MipeGFP transgene for beta-cell labeling.  Total RNA was isolated using TRIzol LS (Invitrogen) and column purification (Zymo Research), according to manufacture's instructions. RNA was DNase-treated (Ambion) and integrity was assessed on the Bioanalyzer (Agilent).    The library was constructed using the Illumina TruSeq DNA sample prep kits A&B, FC-121-1001 and FC-121-1002 with the standard Illumina methods (Bentley 2008).
Experimental Factor: replicate
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