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Basic genetics and patterns of inheritance
Published in Hung N. Winn, Frank A. Chervenak, Roberto Romero, Clinical Maternal-Fetal Medicine Online, 2021
Various banding techniques are used to visualize the chromosomes. The most frequently used method is Giemsa banding or G-banding, which results in a specific pattern of dark and light bands on each chromosome. The older method, quinacrine banding, or Q-banding, produces the same dark and light patterns, but requires the use of a fluorescence microscope and is not used routinely. Reverse banding or R-banding, results in the opposite of the dark and light pattern seen with G-banding; this may be used to better see the ends of the chromosomes. C-banding stains the constitutive heterochromatin, which is near the centromeres and NOR stain visualizes the nucleolar organizing regions of the satellites and stalks of acrocentric chromosomes. For routine karyotype analysis, G-banding is typically used by most laboratories. Routine karyotyping cannot detect gains or losses of cytogenetic material smaller than about 4Mb in size and therefore can miss significant abnormalities.
Clinical Cytogenetics and Testing for Developmental Disabilities
Published in Merlin G. Butler, F. John Meaney, Genetics of Developmental Disabilities, 2019
Joan H. M. Knoll, Linda D. Cooley
Variation (often referred to as a chromosomal heteromorphism or polymorphism) is observed in certain chromosomal regions between homologs of normal chromosomes. It is important to distinguish between normal variation and chromosomal abnormalities. There are two major regions of variation: the first includes size, location, and staining of heterochromatin (genetically inactive chromatin), which is concentrated in distal Yq and the centromeric regions of the other chromosomes, and the second is the morphology of acrocentricshort arms, which have varying amounts of repetitive DNA and copy numbers and activity of ribosomal genes. Additional techniques are utilized to characterize these regions as variants or abnormalities, such as C-banding, which stains heterochromatin, NOR staining, which recognizes active ribosomal genes, and FISH with chromosome-specific centromeric heterochromatin probes or acrocentric short arm probes.
Cytogenetics of Colorectal Cancer
Published in Leonard H. Augenlicht, Cell and Molecular Biology of Colon Cancer, 2019
Three pairs of human chromosomes, Nos. 1,9, and 16, have large blocks of constitutive heterochromatin in the proximal regions of their long arms.86 Quite often, this pericentrometric heterochromatin, as revealed by C-banding, is polymorphic. Atkin and Brito-Babapulle were the first to associate this heterochromatin polymorphism with the development of various malignancies.87 A comparison of C-banding in chromosomes 1,9, and 16 from 128 cancer patients and 111 control individuals revealed that 54.7% of patients were polymorphic in chromosome No. 1, compared to only 35.1% in control. However, the frequency of polymorphism in chromosomes 9 and 16 was not significantly different between the patient and control groups. The polymorphism was primarily in the size differences of C-band region.
An alternative approach for the induction of premature chromosome condensation in human peripheral blood lymphocytes using mitotic Akodon cells
Published in International Journal of Radiation Biology, 2020
Tamizh Selvan Gnana Sekaran, Michelle Ricoul, Patricia Brochard, Cecile Herate, Laure Sabatier
The biodosimetry protocol for PCC of interphase chromosomes from human lymphocytes was initially described by Pantelias (1986). The induction of PCC in PBLs is provoked by fusion with mitotic Chinese hamster ovary (CHO) cells using polyethylene glycol, thus enabling the direct analysis of radiation-induced chromosomal aberrations in non-stimulated G0 lymphocytes. The main advantage of PCC is the rapid quantification of exposure by counting the PCC fragments after Giemsa staining (Pantelias and Maillie 1985a, 1985b; Lamadrid Boada et al. 2013). Various developments of the technique have been published, such as the PCC assay in combination with C-banding (Pantelias et al. 1993) or fluorescence in situ hybridization (FISH), to facilitate the detection of dicentric chromosomes and translocations (Darroudi et al. 1998). More recently, the T/C peptide nucleic acid (PNA) probe, initially developed for metaphase analysis (M’kacher et al. 2014), has been combined with the PCC technique (M’kacher et al. 2015; Karachristou et al. 2015), facilitating dicentric- and ring-chromosome (Dic + R) detection, as well as that of acentric fragments. However, the technique has limitations due to the presence of large interstitial telomeric repeat sequences (ITS) in the large centromeric region of CHO cells.
Chromosomal abnormalities in infertile men with azoospermia and severe oligozoospermia in Qatar and their association with sperm retrieval intracytoplasmic sperm injection outcomes
Published in Arab Journal of Urology, 2018
Mohamed M. Arafa, Ahmad Majzoub, Sami S. AlSaid, Walid El Ansari, Abdulla Al Ansari, Yara Elbardisi, Haitham T. Elbardisi
Cytogenetic investigations were performed on the patient’s chromosomes obtained from peripheral blood lymphocytes, which were cultured in Roswell Park Memorial Institute (RPMI) medium 1640 (Gibco, Invitrogen Carlsbad, CA, USA), phytohemagglutinin (Shanghai Yihua Medical Technology Co., Ltd., Shanghai, China), and foetal bovine serum (Beijing Dingguo Biotechnology, Beijing, China) for 72 h, followed by treatment with 50 μg/mL colcemid. Metaphase chromosome spreads were studied by standard GTG and CBG banding procedures, which included using trypsin and Giemsa for G-banding and barium hydroxide for C-banding. FISH was performed on 30 metaphase chromosome spreads using a mixture of probes specific for X centromere α-satellite (DXZ1) and Y centromere α-satellite (DYZ3; CSP X spectrum green and CSP Y spectrum red; Beijing GP Medical Technologies, Beijing, China), and a chromosome-specific probe for core-binding factor β subunit (CBFB, GLP 16 banding at 16q22, spectrum red; Beijing GP Medical Technologies). Multiplex PCR amplification of nine sequence-tagged site markers was used to detect azoospermia factor (AZF) region for YCMD [19].
Poly (ADP-ribose) polymerase inhibitors sensitize cancer cells to hypofractionated radiotherapy through altered selection of DNA double-strand break repair pathways
Published in International Journal of Radiation Biology, 2022
Yuji Seo, Keisuke Tamari, Yutaka Takahashi, Kazumasa Minami, Shotaro Tatekawa, Fumiaki Isohashi, Osamu Suzuki, Yuichi Akino, Kazuhiko Ogawa
Next, we evaluated the physical rejoining of IR-induced DNA-DSBs by a neutral comet assay (Figure 5(A)). Since the assay for repair foci formation indicated that PARPi reduced the activity of HR, comet tail moments were separately measured in G0- and S-phase-rich cell populations. In both populations, however, the recovery of the tail moments in the combined treatment was not different from that in the IR alone. The results suggested that PJ34 did not inhibit or delay fast repair of DNA-DSBs after IR, although the fidelity of repair was unknown. Unlike the unremarkable result in the physical rejoining of DNA-DSBs, cell cycle profiles showed a striking difference between the combined treatment and IR alone (Figure 5(B)). HCT116 cells were arrested at the G2/M-phases 6–24 h after 10 Gy of IR. The cells were gradually released from the G2/M checkpoint at 24–72 h. Substantial amounts of sub-G1 and populations with eight-fold DNA contents were found at 76 h. The addition of PJ34 or olaparib to IR appeared to augment and prolong the G2/M checkpoint arrests. Populations with 4-fold DNA content remained predominant at 76 h. There was no increase in the sub-G1 population. A G2-phase marker cyclin B1 and an M-phase marker phosphorylated histone H3 were measured in parallel to flow cytometry (Figure 5(C)). After a transient increase at 6 h, cyclin B1 levels were reduced faster in cells treated with the combination of PARPi and IR than in cells treated with IR alone. Phosphorylated histone H3 levels were also lower in cells treated with the combination of PARPi and IR at 24–48 h. These data suggest that PARPi forced cells to evade the G2/M checkpoint and enter into a G0-phase without undergoing a normal mitotic process, which resulted in the cells remaining 4-fold of DNA content. We next measured chromosomal aberrations using a C-banding analysis in HCT116 cells treated with the combination of PARPi and IR compared to cells treated with IR alone. The combined treatment significantly increased the production of dicentric chromosomes compared to IR alone (Figure 5(D,E)). Finally, SA-β-gal staining assays were conducted to explore the mode of death by treatment with PARPi and IR. The addition of PARPi to IR significantly augmented SA-β-gal staining in HCT116 cells (Figure 5(F,G)), in contrast to the flow cytometry results, which only showed modest changes in the sub-G1 population. Taken together, accelerated cellular senescence was likely to be the primary mode of death in HCT116 cells following combined treatment with PARPi and IR.