CGH Pattern Research Articles

Numerical analysis on a viewing angle enhancement of a digital hologram by attaching a pixelated random phase mask.

In a digital hologram, the maximum viewing angle of a computer-generated hologram (CGH) is limited by pixel pitch due to the diffraction grating equation. Since reducing pixel size of display panel is challenging and costly, we propose a method to expand the viewing angle of a digital hologram by attaching an aligned pixelated random phase mask (PRPM) onto the CGH pattern based on analysis of simulation results. By introducing a phase-averaging process to the widely used iterative Fourier transform algorithm, an optimized CGH pattern can be obtained in conjunction with a PRPM. Based on scalar diffraction theory, viewing angle enhancement characteristics were verified by comparing the perspective views of a two-plane hologram using a virtual eye model. In addition, we performed full electromagnetic simulations that included effects due to potential fabrication errors such as misalignment, thickness variation, and internal reflections and diffractions between the CGH and random mask patterns. From the simulation results, by attaching a 1.85 µm-sized pixel pitch PRPM to a 3.7 µm CGH, the viewing angle can be easily expanded almost identical to that of a CGH with 1.85 µm-pixel pitch.

Distinct patterns of complex rearrangements and a mutational signature of microhomeology are frequently observed in PLP1 copy number gain structural variants

BackgroundWe investigated the features of the genomic rearrangements in a cohort of 50 male individuals with proteolipid protein 1 (PLP1) copy number gain events who were ascertained with Pelizaeus-Merzbacher disease (PMD; MIM: 312080). We then compared our new data to previous structural variant mutagenesis studies involving the Xq22 region of the human genome. The aggregate data from 159 sequenced join-points (discontinuous sequences in the reference genome that are joined during the rearrangement process) were studied. Analysis of these data from 150 individuals enabled the spectrum and relative distribution of the underlying genomic mutational signatures to be delineated.MethodsGenomic rearrangements in PMD individuals with PLP1 copy number gain events were investigated by high-density customized array or clinical chromosomal microarray analysis and breakpoint junction sequence analysis.ResultsHigh-density customized array showed that the majority of cases (33/50; ~ 66%) present with single duplications, although complex genomic rearrangements (CGRs) are also frequent (17/50; ~ 34%). Breakpoint mapping to nucleotide resolution revealed further previously unknown structural and sequence complexities, even in single duplications. Meta-analysis of all studied rearrangements that occur at the PLP1 locus showed that single duplications were found in ~ 54% of individuals and that, among all CGR cases, triplication flanked by duplications is the most frequent CGR array CGH pattern observed. Importantly, in ~ 32% of join-points, there is evidence for a mutational signature of microhomeology (highly similar yet imperfect sequence matches).ConclusionsThese data reveal a high frequency of CGRs at the PLP1 locus and support the assertion that replication-based mechanisms are prominent contributors to the formation of CGRs at Xq22. We propose that microhomeology can facilitate template switching, by stabilizing strand annealing of the primer using W-C base complementarity, and is a mutational signature for replicative repair.

Derivation of Genea002 human embryonic stem cell line

The Genea002 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, through ICM outgrowth on inactivated feeders. The line showed pluripotent cell morphology and genomic analysis verified a 46, XY karyotype by CGH and male Allele pattern through STR analysis. Pluripotency of Genea002 was demonstrated with 75% of cells expressing Nanog, 93% Oct4, 83% Tra1-60 and 98% SSEA4, a Pluritest pluripotency score of 24.55, Novelty score of 1.39, teratomas with tissues from all embryonic germ layers and Alkaline Phosphatase activity. The cell line was negative for Mycoplasma and any visible contamination.

Accelerated one-step generation of full-color holographic videos using a color-tunable novel-look-up-table method for holographic three-dimensional television broadcasting.

A color-tunable novel-look-up-table (CT-NLUT) for fast one-step calculation of full-color computer-generated holograms is proposed. The proposed method is composed of four principal fringe patterns (PFPs) such as a baseline, a depth-compensating and two color-compensating PFPs. CGH patterns for one color are calculated by combined use of baseline-PFP and depth-compensating-PFP and from them, those for two other colors are generated by being multiplied by the corresponding color-compensating-PFPs. color-compensating-PFPs compensate for differences in the wavelength between two colors based on their unique achromatic thin-lens properties, enabling transformation of one-color CGH pattern into those for other colors. This color-conversion property of the proposed method enables simultaneous generation of full color-CGH patterns, resulting in a significant reduction of the full color-CGH calculation time. Experimental results with test scenario show that the full color-CGH calculation time of the proposed CT-NLUT has been reduced by 45.10%, compared to the conventional NLUT. It has been further reduced by 96.01% when a data compression algorithm, called temporal redundancy-based NLUT, was used together, which means 25-fold reduction of its full color-CGH calculation time. Successful computational and optical reconstructions of full color-CGH patterns confirm the feasibility of the proposed method.

Computer generated hologram-ROM fabrication and duplication by EBL and UV-NIL

High speed and large capacity read only memory (ROM) is strongly required for the increasing data size of pictures, videos and other files. Computer generated hologram-ROM (CGH-ROM) has the potential possibility to meet these demands and is receiving a lot of attention as the next generation ROM. However, CGH-ROM is difficult to produce in large volume at low cost because it requires a nano-scale precision structure as well as nano-steps. Therefore, in order to fabricate the complex and high-precision pattern cost-effectively, the authors employed ultraviolet nanoimprint lithography (UV-NIL). The three-dimensional (3D) CGH-ROM master mould was fabricated by electron beam lithography (EBL). In this study, the thickness of the residual layer under the CGH pattern was examined. The result showed the target reconstruction image of the CGH-ROM was obtained regardless of a residual layer thickness. Furthermore, the authors have succeeded in fabrication of a CGH-ROM with a 300nm step width and demonstrated its reconstruction.

Sequence based high resolution chromosomal CGH

We aimed to directly align a chromosomal CGH (cCGH) pattern with the gene mapping data by taking advantage of the clustering of the GGCC motif at certain positions in the human genome. The alignment of chromosomal with sequence data was achieved by superimposition of (i) the fluorescence intensity of the sequence specific fluorochrome, Chromomycin A3 (CMA3), (ii) the cCGH fluorescence intensity profile of individual chromosomes and (iii) the GGCC density profile extracted from the Ensembl genome sequence database. The superimposition of these three pieces of information allowed us to precisely localize regions of amplification in the neuroblastoma cell line STA-NB-15. Two prominent cCGH peaks were noted, one at 2p24.3, the position 15.4 mega base (Mb), and the other at 2p23.2, 29.51 Mb. FISH and high resolution array CGH (aCGH) experiments disclosed an amplification of MYCN (16 Mb) and ALK (29.2–29.9 Mb), thus confirming the cCGH data. The combined visualization of sequence information and cCGH data drastically improves the resolution of the method to less than 2 Mb.

Frequent Molecular Aberrations in Normal Karyotype (NC) Precursor B-Cell Acute Lymphoblastic Leukemia (pB-ALL) Detected by Comparative Genomic Hybridization.

The prognosis of adult NC pB-ALL has not improved over the last decade mainly because separation into distinct molecular subsets has been lacking. We conducted a proof-of-principle experiment to screen the genome of blasts from adult NC pB-ALL patients for novel genomic alterations using 19,000 bacterial artificial chromosomes (BAC) by array CGH and verified our results with gene expression profiling (GEP, Agilent oligonucleotide expression arrays) and fluorescent in situ hybridization (FISH) using the same BAC probes or commercially available probes if available. The median age at diagnosis of the 6 males and 4 females was 55 years; all were treated with a 5-drug induction regimen followed by intensive consolidation and maintenance regimens. Five achieved CR of whom 3 relapsed with complex karyotype; samples were available on 2 of the patients at relapse. Samples with <60% blasts (2 at diagnosis, 1 at relapse) were enriched for >95% purity by sorting according to their immunophenotype pattern. Only one case had a normal CGH pattern. A decreased copy number of p16 was found in 4 of the 10 diagnostic samples; 3 on whom mRNA was available demonstrated a decreased p16 mRNA expression by GEP. In addition, a loss at 9p21.2 and a loss and a gain at 8q21.2 were detected in one sample each; those were verified by FISH. Copy number losses were also observed across multiple samples at 1p34–1p36, 4p16, 5p15.33, 8q24, 9q34, 10q26, 11q12–11q13, 12q24, 14q32, 17p, 19p13.3, 22q11, and 22q13. Copy number gains across multiple samples were observed at 15q11.2 and 18q21. Those are currently being verified by FISH. Comparison of samples at diagnosis and at relapse revealed a disparate molecular pattern. In summary, enriching samples by sorting from NC pB-ALL to be studied by CGH is feasible; normal chromosome pattern does not correlate with normal molecular pattern; and diagnostic and relapsed samples have a different molecular pattern. Our work on 10 patient samples at diagnosis and 2 at relapse demonstrates the feasibility to detect specific molecular aberrations in NC pB-ALL. Since attaining good quality metaphase cells in ALL is often suboptimal, this work suggests that array CGH can complement conventional cytogenetic analysis to better characterize NC pB-ALL by identifying cryptic copy number changes. Work on a larger number of samples is warranted to determine the prognostic significance of the aberrations detected herein.

P53 mutations as an identification marker for the clonal origin of bladder tumors and its recurrences

The clonality of synchronous and metachronous bladder tumors has been studied for years with controversial results. Some recent studies support the 'polyclonal origin' hypothesis, i.e. that independently transformed different tumor cell clones exist in the same bladder cancer patient and arise from the field cancerogenisation affecting the entire bladder urothelium by environmental mutagens. Others could demonstrate a monoclonal origin of primary bladder tumors and its recurrences due to a single genetically transformed cell clone spread through the urinary system. With increasing understanding of the clonal origin of bladder tumors and recurrences, clonality markers might contribute to an early and accurate prediction of tumor recurrence and progression. We used p53 mutations as an identification marker permitting the prediction of clonality in bladder tumors and its recurrences. Primary tumors (n=33) and recurrences (n=63) were screened by direct genomic sequencing the p53 mutation hot spot region, exons 5-8. P53 mutations occurred in 12% in our cohort, predominantly in higher malignant (>or=G2), invasive (>or=T1) tumor samples. We were able to demonstrate intratumoral heterogeneity regarding the p53 status and that recurrences may occur from genetically unrelated primary tumor sites. Some of our results argue for a polyclonal origin of synchronous and metachronous bladder tumors possibly due to the field effect in bladder carcinogenesis. Evidence for a monoclonal origin was found in two cases: one case with a high malignant primary tumor and 3 metachronous recurrences, all of them harbouring the same exon 8 mutation found in the primary tumor; one case with identical mutations of exon 8 in the primary and one recurrent tumor. For further implications concerning clonality of recurrent bladder tumors, p53 status should be combined with a broader range of markers such as CGH and LOH pattern.