human testis total protein  (AMS Biotechnology)

Journal: BMC Evolutionary Biology

Article Title: Primate-specific spliced PMCHL RNAs are non-protein coding in human and macaque tissues

doi: 10.1186/1471-2148-8-330

Figure Lengend Snippet: Identification of spliced PMCHL transcripts . (A) PMCHL exon/intron structure as deduced from previous and present transcript sequences. Primer positions are indicated. (B) Identification of alternative spliced transcripts by PCR and sequencing. For human testis, transcripts harbouring exons 1 and 6a were amplified using primer pair 3–9/3–30, and transcripts harbouring exons 1 and 6b were amplified using primer pair 3–9/3–353, followed by 3–10/3–353. For human fetal brain, PCR primer pairs were 3–7/3–30, followed by 3–8/3–27. Documented ESTs [GenBank: AI203691 ; EMBL: BX091674 ; GenBank: AA724728 ; GenBank: BG184695 ] corresponding to spliced transcripts are represented. The number of independent clones for each transcript identified in our previous and present studies are indicated in brackets. The 3b splice donor site differs in PMCHL1 (named 3b 1 ) and PMCHL2 (named 3b 2 ). Two 5b splice acceptor sites (indicated by t/b), separated by four nucleotides, were identified. GenBank accession numbers of transcripts are: [GenBank: EU921424 , GenBank: EU921425 , GenBank: EU921426 , GenBank: EU921427 , GenBank: EU921428 , GenBank: EU921429 , GenBank: EU921430 , GenBank: EU921431 , GenBank: EU921432 , GenBank: EU921433 , GenBank: EU921434 , GenBank: EU921435 , GenBank: EU938381 ]. (C) RT-PCR and Southern blot analysis of spliced transcripts in human and macaque adult testis, prefrontal cortex (CX) and cerebellum (CB). Spliced transcripts were detected only in human testis. PCR amplification was with primer pair 3–7/3–30. Molecular weights are indicated. M, size markers; RT, reverse transcribed; NRT, non-reverse transcribed.

Article Snippet: Human adult testis total proteins were purchased from BioChain/Cliniscience (France).

Techniques: Sequencing, Amplification, Clone Assay, Reverse Transcription Polymerase Chain Reaction, Southern Blot, Reverse Transcription

Journal: BMC Evolutionary Biology

Article Title: Primate-specific spliced PMCHL RNAs are non-protein coding in human and macaque tissues

doi: 10.1186/1471-2148-8-330

Figure Lengend Snippet: Analysis of the coding potential of VMCH-p8 and its variants in human and macaque tissues . (A) Schematic representation of PMCHL ORF1, encoding the putative VMCH-p8 protein, present on unspliced RNA, and its variants, ORF1a and ORF1b, present on spliced RNAs. (B) Sequence comparison of PMCHL1 and PMCHL2 ORF1, ORF1a and ORF1b. The ORF1 epitope is underlined. The putative nuclear localisation signal (NLS) is shaded in grey. The variable C-terminal parts are boxed. (C) Sequence comparisons of PMCHL1 ORF1, ORF1a and ORF1b in Homo sapiens ( HSA ), Pan troglodytes ( PTR ), Pongo pygmaeus ( PPY ) and Macaca fascicularis ( MFA ). The variable C-terminal parts are boxed. (D) Specificity of the anti-VMCH-p8 antiserum. In Western blots, immune anti-VMCH-p8 serum detects the recombinant GST-VMCH-p8 protein (12.5 ng) migrating at 34 kD (arrow). (E) Western blot analysis of the expression of VMCH-p8 and its variants in vivo in human and macaque tissues. GST-VMCH-p8 (12.5 ng), human adult testis proteins (25 μg), new-born (NB) and fetal (F) hippocampus (HPC) and prefrontal cortex proteins (CX) (80 μg each), and macaque supplementary motor area (SMA), cerebellum (CB), prefrontal cortex (CX) and visual area (VA) proteins (80 μg each) were analysed using the anti-VMCH-p8 antiserum. The arrow points to GST-VMCH-p8 migrating at 34 kD.

Article Snippet: Human adult testis total proteins were purchased from BioChain/Cliniscience (France).

Techniques: Sequencing, Comparison, Western Blot, Recombinant, Expressing, In Vivo