Spatial / Single Cell Transcriptomics, Gene Ontology, Protein

is reliable and accurate [10, 15]. In light of these findings, a targeted whole transcriptome kit was recently developed (AmpliSeq, Life Technologies AmpliSeqTM technology). This kit is designed for targeted amplification of over 20,000 distinct human RNA targets simultaneously in a single primer pool.

염기시열 분석법(custom targeted RNA-seq) 방법인 AmpliSeq을 개발하여 이를 소개하였다. 현재까지 NGS을 이용한 RNA-seq은 전체 전사체(whole transcriptome) 수준으로 이용되었으나 모든 유전자를 확인함에 따른 비용 증가와 유전자 간의 균일한 분석이 어렵다는 점에서 단점이 있었다. 이를 해결하기 위해 Ilumina에서는 유전자 세트를 특정하여 분석할 수 있는 상용 표적 RNA 염기시열 분석법인 AmpliSeq을 개발하였다..
AmpliSeq은 라이브러리 제작, 데이터 생성 및 분석의 과정을 간소화하고 간편화하였다. 최소 1 ng의 RNA로부터 80여 개 유전자 발현을 측정할 수 있다.3435

Spatial transcriptomics

Methods
Microdissection
(simply) scrapoing regions of interest off a slide using a small needle

Single cell transcriptomics

Experimental Steps

	steps	method
↓	Cell isolation	FACs
	Lysate formation
	amplification through reverse transcription and	RNA-seq
	quantification of expression levels

Data analysis

Differential expression		Clustering	Dimensionality reduction
Gene ontology enrichment[edit]	Pathway enrichment analysis: Compared to other pathways, are there more genes in the pathway differentially expressed (up-regulated/downregulated) ? To reduce data dimensionality by arranging genes into pathways. · Helps interpret the data in the context of biological processes, pathways and networks.	k-means clustering, hierarchical clustering	Principal component analysis[edit] The most frequently used technique is PCA, which identifies the directions of largest variance principal components and transforms the data so that the first principal component has the largest possible variance, and successive principle components in turn each have the highest variance possible while remaining orthogonal to the preceding components. The contribution each gene makes to each component is used to infer which genes are contributing the most to variance in the population and are involved in differentiating different subpopulations.[19] PCA transform the large scale of data (e.g., 100 sample by 100 proteins into a matrix with 100 sample by a few numbers of columns [PCs]).

Gene ontology

• Database
  • GO website[?] in a number of formats, or can be accessed online using the GO browser AmiGO.
• EXAMPle
  • id: GO:0000016
  • name: lactase activity
  • ontology: molecular_function
  • def: “Catalysis of the reaction: lactose + H2O=D-glucose + D-galactose.” [EC:3.2.1.108]
  • synonym: “lactase-phlorizin hydrolase activity” BROAD [EC:3.2.1.108]
  • synonym: “lactose galactohydrolase activity” EXACT [EC:3.2.1.108]
  • xref: EC:3.2.1.108
  • xref: MetaCyc:LACTASE-RXN
  • xref: Reactome:20536
  • is_a: GO:0004553 ! hydrolase activity, hydrolyzing O-glycosyl compounds

[Protein]

Code

Gene name → UniProKB/Swiss-Prot	UniProKB/Swiss-Prot → Gene name
https://www.uniprot.org/id-mapping 에 gene name을 입력하시고요. → from 에서 uniprot -> gene name을 선택하시고, → to는 uniprotkb 에서 UniProKB/Swiss-Prot 선택 → toxonomy 입력란이 추가로 생겨요. 거기에 homo sapiens 입력하시고 결과를 만드세요. 그후에 결과를 다운로드 받으실때 엑셀 파일 선택	https://www.uniprot.org/id-mapping From: UniProt에서 UniProtKB AC/ID 선택. To : UniProt에서 Gene name 선택

Structure of protein

	Start	End
	N terminal free amine group (-NH2)	C (carboxyl) terminal terminated by a free carboxyl group (-COOH)
translation	When the protein is translated from messenger RNA, it is created from N-terminus to C-terminus.
function	Signal for exiting ribosome, signal for half-life	Signal for ER retention
Posttranslational modification	various	various

Species of protein

• fibrillar protein any of the generally insoluble proteins that comprise the principal structural proteins of the body, e.g., collagens, elastins, keratin, actin, and myosin.

Here we provide a detailed characterization of Aβ42cc protofibrils. Like wild type Aβ they appear as smooth rod-like particles with a diameter of 3.1 (±0.2) nm and typical lengths in the range 60 to 220 nm
 
Protofibrils are transient structures observed during in vitro formation of mature amyloid fibrils

Figure labels (Aβ42cc amyloid aggregation, evidence-only):

Monomers → Oligomers → Protofibrils → Mature fibrils
N ─────► C  (each species labeled with N- and C-termini and connecting arrows)

Turnover of proteins

• https://www.quora.com/Do-mice-have-protein-turnover-rates-that-are-different-than-that-of-humans
• (?eb et al. 2019, PMID 31357232) longer lifetimes of proteins in comparison with their mRNA counterparts, with a mean lifetime of roughly 40 min.

Uncertain Spans

location	text/status	reason
AmpliSeq Korean paragraph	trailing token 3435	Looks like a citation marker pair; visible at the end of the Korean paragraph but its exact meaning (citation IDs vs. concatenated marks) is not separable from this photo.
Single cell Experimental Steps table	left-most column showing only ↓	The table has a thin left column that displays a downward arrow spanning the rows; treated as a flow indicator.
Gene ontology Database bullet	trailing icon after GO website	A small icon/letter follows GO website; it cannot be resolved at this resolution.
Turnover citation	(?eb et al. 2019, PMID 31357232)	First-author surname is partly clipped at the bottom edge; only eb et al. is visible.