TY - JOUR

T1 - Characterization of axolotl lampbrush chromosomes by fluorescence in situ hybridization and immunostaining

AU - Keinath, Melissa C.

AU - Davidian, Asya

AU - Timoshevskiy, Vladimir

AU - Timoshevskaya, Nataliya

AU - Gall, Joseph G.

N1 - Funding Information: This work was supported by the National Institute of General Medical Sciences of the National Institutes of Health (grant number R01 GM33397 to J.G.G). J. G.G. is American Cancer Society Professor of Developmental Genetics. The BAC clones were funded through grants to S Randal Voss from the National Institutes of Health (grant numbers R24 RR016344 , R24 OD010435 ) and the Department of Defense (grant number W911NF1110475 ). Animals were obtained from the Ambystoma Genetic Stock Center funded by National Institutes of Health grant number P40OD019794 to S.R.V. Publisher Copyright: © 2021 Copyright: Copyright 2021 Elsevier B.V., All rights reserved.

PY - 2021/4/15

Y1 - 2021/4/15

N2 - The lampbrush chromosomes (LBCs) in oocytes of the Mexican axolotl (Ambystoma mexicanum) were identified some time ago by their relative lengths and predicted centromeres, but they have never been associated completely with the mitotic karyotype, linkage maps or genome assembly. We identified 9 of the axolotl LBCs using RNAseq to identify actively transcribed genes and 13 BAC (bacterial artificial clone) probes containing pieces of active genes. Using read coverage analysis to find candidate centromere sequences, we developed a centromere probe that localizes to all 14 centromeres. Measurements of relative LBC arm lengths and polymerase III localization patterns enabled us to identify all LBCs. This study presents a relatively simple and reliable way to identify each axolotl LBC cytologically and to anchor chromosome-length sequences (from the axolotl genome assembly) to the physical LBCs by immunostaining and fluorescence in situ hybridization. Our data will facilitate a more detailed transcription analysis of individual LBC loops.

AB - The lampbrush chromosomes (LBCs) in oocytes of the Mexican axolotl (Ambystoma mexicanum) were identified some time ago by their relative lengths and predicted centromeres, but they have never been associated completely with the mitotic karyotype, linkage maps or genome assembly. We identified 9 of the axolotl LBCs using RNAseq to identify actively transcribed genes and 13 BAC (bacterial artificial clone) probes containing pieces of active genes. Using read coverage analysis to find candidate centromere sequences, we developed a centromere probe that localizes to all 14 centromeres. Measurements of relative LBC arm lengths and polymerase III localization patterns enabled us to identify all LBCs. This study presents a relatively simple and reliable way to identify each axolotl LBC cytologically and to anchor chromosome-length sequences (from the axolotl genome assembly) to the physical LBCs by immunostaining and fluorescence in situ hybridization. Our data will facilitate a more detailed transcription analysis of individual LBC loops.

KW - Ambystoma mexicanum

KW - Axolotl

KW - BAC FISH

KW - Centromere

KW - Lampbrush chromosomes

KW - Ambystoma mexicanum/genetics

KW - In Situ Hybridization, Fluorescence

KW - Chromosome Mapping

KW - Chromosomes, Artificial, Bacterial/genetics

KW - Animals

KW - Oocytes/growth & development

KW - Transcription, Genetic

KW - Chromosomes/genetics

KW - Centromere/genetics

UR - http://www.scopus.com/inward/record.url?scp=85102040150&partnerID=8YFLogxK

UR - https://www.mendeley.com/catalogue/37e77ed9-e013-3819-a92a-16d6afb68ea3/

U2 - 10.1016/j.yexcr.2021.112523

DO - 10.1016/j.yexcr.2021.112523

M3 - Article

C2 - 33675804

AN - SCOPUS:85102040150

VL - 401

JO - Experimental Cell Research

JF - Experimental Cell Research

SN - 0014-4827

IS - 2

M1 - 112523

ER -