WGA analysis and aCGH of single sperm.

<p><b>a</b>. Gel electrophoresis of WGA products from single human sperm. Ct: cycle threshold as determined by Real-Time PCR with SYBR Green. <b>b</b>. Real-Time PCR of the same WGA products. The difference in fluorescence (fluorescence at the end of each cycle minus baseline fluorescence level) is plotted against the cycle number. Hybridization failed for samples with a cycle threshold >10. <b>c</b>. Molecular karyotype of a normal 23,X sperm co-hybridized with female (upper panel) and male reference (lower panel). <b>d</b>. Molecular karyotype of a normal 23,Y sperm co-hybridized with a female (upper panel) and a male reference (lower panel).</p

Subject characteristics and summary of aCGH results on single sperm.

<p>Subject characteristics and summary of aCGH results on single sperm.</p

Flow diagram of the protocol developed for aCGH on single human sperm.

<p>Flow diagram of the protocol developed for aCGH on single human sperm.</p

Detection and localization of HPV in human sperm.

<p><b>a</b>. Fluorescence in situ hybridization (fluorescence microscope) for HPV DNA on sperm from a patient with HPV16 in semen. Infected and noninfected sperm are shown. Red: HPV DNA (Texas red); blue: nuclear staining (DAPI). <b>b</b>. Immunofluorescence (confocal fluorescence microscope) for HPV16 capsid protein L1 on sperm from a control (left) and a patient with HPV16 in semen (right). Upper panel, L1 antibody; central panel, L1 antibody and Pisum Sativum (acrosome); lower panel, L1 antibody and Pisum Sativum after induction of the acrosome reaction. Red: HPV16 L1; green: Pisum Sativum; blue: nuclear staining (DAPI). <b>c</b>. PCR for HPV E7 gene from sperm DNA. Lane M: DNA marker (100 bp); 1: negative control (no template); 2: positive control (sperm transfected with recombinant plasmid pIRES2-AcGFP1-E6E7); 3: sperm from a patient with HPV16 in semen; 4: sperm from a control subject.</p

Oocytes penetrated by sperm transfected with HPV E6/E7 express HPV E6/E7 genes.

<p><b>a</b>. PCR for E6 gene from single oocyte penetrated by transfected sperm. Lane M: DNA marker (100 bp); 1: positive control (pIRES2-AcGFP1-E6E7 plasmid); 2: single oocyte penetrated by sperm transfected with recombinant plasmid; 3: negative control (no template); 4: single oocyte penetrated by sperm transfected only with Lipofectamine 2000. <b>b</b>. GFP-E6 expression in oocyte penetrated by control sperm. <b>c</b>. GFP-E6 expression in oocyte penetrated by transfected sperm. <b>d</b>. RT-PCR for E6 gene from single oocyte penetrated by transfected sperm. Lane M: DNA markers (100 bp); 1: negative control (no template for Reverse Transcription); 2: negative control (no template for PCR); 3: single oocyte with green fluorescence penetrated by transfected sperm; 4: single oocyte without green fluorescence penetrated by control sperm.</p