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Animal Genetics Mind Map (Zhongnong Genetics Class Assignment)
Content; Molecular biology basis of inheritance, transmission of genetic information, gene expression and regulation, changes in genetic material, genome and genetic engineering; continuously updated!
Edited at 2024-03-24 20:43:46
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Animal Genetics Mind Map (Zhongnong Genetics Class Assignment)
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Animal genetics guide copy 2
Molecular Biological Basis of Heredity
carrier of genetic information
The process of discovery of genetic material
In 1928, Frederick Griffth's bacterial transformation experiment demonstrated that R-type bacteria received transformation factors from dead S-type bacteria and transformed into S-type bacteria, gaining pathogenicity.
In 1944, Oswald Avery, et al. analyzed genetic material and proved that DNA is the transformation factor that transforms R-type bacteria into S-type bacteria, and DNA is the carrier of genetic information (forward basis and reverse basis)
The phage infection experiment once again proved that DNA is genetic material (scientific conclusions need to be repeatedly verified)
It was discovered that the genetic material of tobacco mosaic virus is RNA, and other carriers of genetic material were discovered - RNA (there are always "exceptions" to the general rule)
Basis for identification of genetic material (i.e. basic characteristics of genetic material)
Can control metabolic processes and expression of traits
Correspondence between genes and phenotypes
Able to reproduce itself, allowing previous and subsequent generations to maintain a certain degree of continuity
Phage infection experiment
can cause heritable variation
Infection with plant viruses
Infection with animal viruses - virus control/complete cure
Indirect evidence for DNA as the primary genetic material:
Content: DNA content is constant. The DNA content in gametes is half that of somatic cells, and the DNA content in polyploids is doubled
Metabolism: DNA molecule metabolism is relatively stable
Mutation: The most effective wavelength when ultraviolet light induces mutation is consistent with the ultraviolet spectrum absorbed by DNA, proving that genetic mutation is closely related to the variation of DNA molecules.
Distribution: DNA is shared by all biological chromosomes (some viruses do not contain DNA and use RNA as genetic material)
Methods to verify the genetic basis of traits
Correspondence between genes and phenotypes—relevance, necessity, and sufficiency
Discovery of chicken green-shell egg gene
Verification of function of Drosophila purple eye gene
molecular structure of nucleic acids
nucleic acid
Nucleotides are the basic structural units of DNA and RNA, formed by the dehydration condensation of pentose sugars with bases and phosphates
DNA structure and biological significance
Primary structure of DNA - chemical composition Features:
The primary structure of DNA refers to the linking method and arrangement order of the four nucleotides in the DNA molecule, usually represented by the base sequence.
Chargaff's equivalent law: [A]=[T], [G]=[C], A G=C T
The first law - double-stranded DNA, applies to partial and complete genomes
The second law - single-stranded DNA, applies to the complete genome of double-stranded DNA
The base sequence of DNA molecules is arranged in extremely diverse ways, and changes in the base sequence may cause changes in genetic information.
Secondary structure of DNA - Watson-Crick DNA double helix structure key points:
Right-handed helix, two complementary strands antiparallel
The phosphate group and deoxyribose are on the outside, and the bases are on the inside of the helix.
The two strands pair up complementary by hydrogen bonds
The diameter of the double helix is 2nm, the pitch is 3.32nm, and each helix has 10.5 base pairs.
Major grooves and minor grooves appear alternately on the surface of the helix, and proteins recognize bases through these two grooves.
Polymorphism of DNA structure: (the meaning of DNA conformation)
There are 7 known DNA double helix conformations. A, B, C, D, E, and T types are right-handed helices, the W-C model is the B conformation, and Z-DNA is a left-handed helix.
Advanced structure of DNA - physical conformation
The higher-order structure of DNA refers to the specific spatial structure formed by the further twisting and coiling of the DNA double helix.
Superhelical structure is the main form of higher-order structure of DNA, which can be divided into two types: negative superhelix (natural double helix DNA) and positive superhelix.
DNA and chromosomes
Nucleosomes are the basic structural units of chromatin and are composed of DNA and histones.
DNA makes up 30-40% of chromatin mass
The DNA sequence of the special structure of chromosomes
Euchromatin: A lightly stained segment of chromatin threads
Heterochromatin: A deeply stained segment of chromatin strands
Constitutive heterochromatin: mainly satellite DNA, such as centromeres
Facultative heterochromatin: found anywhere on a chromosome, such as the X chromosome in mammals
RNA classification and structural characteristics
Both prokaryotes and eukaryotes contain many different RNA molecules, the most important of which are:
Messenger RNA - transmits genetic information, accounting for 5-10% of total RNA
Transfer RNA - small molecular weight RNA, transport amino acids, recognition codons, clover-type secondary structure, accounting for 10-15% of the total RNA
Ribosomal RNA - composed of large and small subunits, the site of protein synthesis and assembly, accounting for 75-80% of total RNA
Coding RNA and non-coding RNA
Coding RNA: RNA that directs the translation of proteins
Non-coding RNA: RNA that does not code for proteins is transcribed from the genome, but is not translated into proteins and can perform its biological functions at the RNA level. Including rRNA, tRNA, snRNA, snoRNA and microRNA and other RNAs with known functions, as well as RNAs with unknown functions
Examples: small RNA (micro-RNA), long non-coding RNA
3D genome sequencing and T2T sequencing
Gene
Origin of the concept of genes
Mendel, hereditary factors - the abstract concept of genes
Johnson, Gene replaces genetic factors, genotype and phenotype - the noun system of genes
Morgan, genes are located on chromosomes - the material carriers of genes
Avery, Hirsch and Chase, the chemical essence of genes is DNA - the material basis of genes
Watson and Crick, proposed the double helix model of DNA - how genetic information is transmitted
The development of genetic content
Jumping gene: refers to a gene whose position can be moved on the chromosome, for example, a transposon.
Operator: Many functionally related genes are connected in a string and controlled by a common control region for transcription, including the entire DNA sequence of structural genes and regulatory genes.
Broken genes: The coding sequences of eukaryotic genes are often discontinuous. They are separated by some non-coding sequences, forming a broken gene structure.
Overlapping genes: refers to the existence of shared DNA sequences between two genes.
Pseudogene: A gene that has lost its original function. An inactivated gene that has a similar sequence to a normal functional gene but cannot synthesize functional proteins due to mutations.
Fusion gene: Transcription starts from a DNA sequence encoding one protein and proceeds to another gene encoding a completely different protein.
Non-coding genes: genes that do not contain instructions for making proteins and do not function in the form of proteins
An exon encoding a protein can cross chromosomal boundaries by combining with an exon elsewhere in the gene.
apparent genetics
Definition of gene (2015): Gene: It is a region associated with regulation, transcription or function that can be found in the genome sequence and corresponds to a genetic unit.
Classification of genes (according to functional form)
Genes encoding proteins with transcription and translation functions
Non-coding RNA genes, genes that only have transcribed products without translation
Genes that are not transcribed exist in the form of DNA and regulate gene expression.
General structure of eukaryotic coding genes
The region translated into a peptide chain - start codon to stop codon
The region transcribed into mRNA—transcription start site to transcription termination site, including 5’UTR, coding region, and 3’UTR
Region involved in the transcriptional regulatory process – 5’ flank to 3’ flank
Main elements in eukaryotic gene structure
Introns and exons: RNA splicing signals, the GT-AG rule
Transcription start site: the first base on the template when transcription begins (1)
Promoter: the site where RNA polymerase recognizes and binds (-100bp)
Enhancer: improves the transcription efficiency of genes. Its effect has nothing to do with its position, direction and distance from the gene. It is tissue-specific and cell-specific.
Silencer: Binds to a specific protein to inhibit gene transcription
Insulator: Located between the promoter and regulatory elements, it prevents the regulatory elements from functioning and its effect is directional.
transmission of genetic information
DNA replication
Definition: The process of synthesizing new progeny DNA molecules with the same structure as the parent template using the parent DNA molecule as a template.
Basic laws of DNA replication
Replication origins and replicators
Related enzymes and proteins involved in DNA replication
DNA polymerase - polymerization, exo-functionality
prokaryotic DNA polymerase
DNA polymerase I (DNA proofreading repair), II (DNA damage repair)
DNA polymerase I has 5’-3’ exonuclease activity and excises the 5’-RNA primer.
DNA polymerase III is the main enzyme for bacterial DNA replication
Characteristics of DNA polymerase III: 5'-3' polymerase activity, catalyzes the addition of dNTP to the 3'-OH end of the growing DNA chain, and does not have the ability to directly initiate DNA synthesis; 3'-5' exonuclease activity , plays a proofreading function; does not have 5'-3' exonuclease activity.
eukaryotic DNA polymerase
There are five types of DNA polymerases in eukaryotes: α, β, γ, δ and ε. DNA polymerase
δ is considered to be the main enzyme catalyzing DNA replication in eukaryotes.
Helicase – Unwinds the double strands of DNA.
Single-stranded binding protein - protects DNA from hydrolysis and from returning to double strands.
Topoisomerase - I - eliminates positive supercoils (compacts), II - restores negative supercoils (relaxes).
Prima enzyme - RNA polymerase, catalyzes the synthesis of RNA primers (11-12nt).
DNA polymerase requires a 3' free hydroxyl group, but RNA polymerase does not. Eukaryotic DNA polymerase α has initiase activity.
DNA ligase – ligates Okazaki fragments but not single-stranded DNA molecules
General process of DNA replication
Initiation of DNA replication, replication fork
Elongation of DNA replication, replication fork advancement, lagging strand formation
Termination of DNA replication: RNA primer excision, gap filling, Okazaki fragment ligation, and restoration of supercoiling. Unidirectional replication of circular DNA terminates near the origin of replication; the replication endpoint of bidirectional replication of linear DNA and circular DNA is not fixed.
Characteristics of eukaryotic DNA replication
Eukaryotic DNA replication occurs in a specific phase of the cell cycle - the synthesis phase (S) of the interphase of cell division. Generally, the second round of replication occurs only after the first round of replication is completed; in prokaryotes during the entire cell growth process All can carry out DNA replication, and the origin of replication can initiate replication continuously.
Cells can be divided into three categories based on their ability to divide:
Cyclic cells always maintain active division ability and continuously enter the cell cycle cycle, such as hematopoietic stem cells, stem cells of epidermis and gastrointestinal mucosa epithelium.
The group of cells that are not proliferating (G0 phase cells) are differentiated cells that perform specific functions. They are usually in the G0 phase, so they are also called G0 phase cells. Under certain stimulation, these cells re-enter the cell cycle. Such as liver cells, renal tubular epithelial cells, cardiomyocytes, and thyroid follicular epithelial cells. After partial hepatectomy, the remaining liver cells divide rapidly.
Terminal differentiated cells, which have lost the ability to divide, are also called terminal cells, such as mature red blood cells, nerve cells and other highly differentiated cells.
Eukaryotes have many replication origins, which are not activated simultaneously during S phase. At different developmental stages, the number of replication origins and replicon size of eukaryotes will change.
The DNA replication rate (number of bases copied per minute) of eukaryotes is slower than that of prokaryotes. However, eukaryotes have multiple replicons, so the replication speed of the entire chromosome is not slower than that of prokaryotes.
Reassembly of eukaryotic nucleosomes. Eukaryotic DNA replication is coupled with nucleosome assembly. The replication fork synthesizes new DNA strands and assembles nucleosomes at the same time. Old histone modifications can be transferred to new nucleosomes. The DNA of prokaryotes is naked, with no histones bound to it and no nucleosome structure.
The ends of eukaryotic chromosomes - telomeres: are composed of short sequence tandem repeats and telomere-binding proteins, which protect the chromosome structure. The 3' single-stranded end is folded into the double-stranded repeat sequence to form a loop to protect the chromosome ends. Telomeres, centromeres and origins of replication are the three major elements that keep chromosomes intact and stable.
Replication of DNA at the ends of eukaryotic chromosomes: Linear DNA molecules, after the end primer is removed, due to the absence of free 3'-OH, the 5' end cannot be synthesized and causes deletions.
The significance of telomere repair
The relationship between DNA replication and inheritance:
transmission of genetic information
cell proliferation, renewal
Occurrence and Correction of Variations
Form the basis for genetic diversity, evolution or new traits
Trait selection or disease treatment targets
gene transcription
Using a single strand of DNA as a template and four ribonucleotides as raw materials, the process of synthesizing an RNA chain under the catalysis of RNA polymerase
Similarities and Differences between Transcription and Replication
Similarities between Transcription and Replication
Catalysis by polymerase
Using DNA as a template
Synthesized according to the principle of complementary base pairing
Generate new strands along the 5’ to 3’ direction
Difference Between Transcription and Replication
Transcription only occurs in part of the region
During transcription, only one strand serves as a template, which is called the template strand or antisense strand, while the other is called the sense strand or coding strand.
Transcription initiation does not require the participation of primers, RNA polymerase does not require 3’ free hydroxyl groups, and RNA chain synthesis is continuous.
The substrates for transcription are four ribonucleoside triphosphates (rNTPs), namely ATP, GTP, CTP and UTP
During transcription, the DNA-RNA hybrid double-stranded molecule is unstable. The single-stranded RNA continuously detaches from the template strand during the extension process, and the template DNA returns to the double-stranded state.
The primary transcripts generated by transcription of eukaryotic genes generally need to be processed to become mature RNA molecules and have biological functions.
RNA polymerase
E. coli RNA polymerase
There is only one RNA polymerase in E. coli that is responsible for the synthesis of all mRNA, rRNA and tRNA. It is a complex enzyme composed of 5 subunits (α2 ββ’σ). The four subunits of α2ββ' constitute the core enzyme. The core enzyme and the σ subunit constitute the holoenzyme. The core enzyme has polymerization activity. The σ subunit has no catalytic activity, but can recognize the promoter and form a stable initiation complex with DNA. Involved in the initiation of transcription.
Promoter (prokaryotic): It is a component of a gene. It is a region on the DNA molecule that can bind to RNA polymerase and form a transcription initiation complex to control gene transcription and expression.
eukaryotic RNA polymerase
There are three types of RNA polymerase responsible for transcription in eukaryotic cells: RNA polymerase I, II and III, which are located in different parts of the cell. RNA polymerase II is the main enzyme for mRNA synthesis, catalyzing the synthesis of mRNA and small nuclear RNAs (such as snRNA, microRNA).
Promoter (eukaryotic): The promoter is a component of the gene, like a "switch" that determines the activity of the gene and controls the start time and degree of gene expression (transcription). Promoters themselves do not control gene activity alone, but rather by binding to such proteins called transcription factors (TFs).
General process of gene transcription
Transcription initiation
Core enzyme factor search promoter
Melted by RNA polymerase
The holoenzyme moves to the transcription start site and synthesizes the first rNTP of the RNA chain.
The factor dissociates, the affinity of the core enzyme to the template decreases, and it moves on the DNA
extension of transcription
After transcription is initiated, the σ factor has dissociated and the elongation process is catalyzed by the core enzyme
RNA polymerase moves in the 3’-5’ direction of the template DNA strand, and the newly synthesized RNA chain itself extends in the 5’-3’ direction.
The region covered by the core enzyme unwinds the DNA double strands
The hybrid strand region of RNA and DNA pairing is about 12bp
termination of transcription
Terminator: It is a sequence that exists in the primary transcript, generally located downstream of the poly(A) site, and is about a few hundred bases in length.
Strong terminator: It has a palindromic structure and is rich in GC sequences, which is difficult to unravel and prevents RNA polymerase from moving forward. There are oligomeric Us with multiple nucleotides at the 3' end, forming A-U pairing with the template, making the RNA easy to release. Transcription is completed without the help of other protein factors.
Weak terminator: There are also palindromic sequences with little G-C content in the hairpin structure and no oligomeric U at the 3' end. It causes the polymerase to pause. Without the help of other protein factors, the polymerase will continue to "read through" past the terminator. Transcription is terminated only when the protein factor ρ is present.
Rho factor-dependent transcription termination mechanism
Characteristics of eukaryotic DNA transcription
RNA polymerase
There are three types of RNA polymerase in eukaryotes, each of which transcribes different RNAs; there is only one type of RNA polymerase in prokaryotes, which catalyzes the synthesis of all RNA.
initiation of transcription
Eukaryotic RNA polymerase cannot independently transcribe RNA. It must have a promoter protein (transcription factor) bound to the promoter in order to bind to the promoter. The regulatory protein binding sites include TATA box, CAT box, GC box, enhancer sequence, etc.
RNA transport and processing
After eukaryotic RNA is transcribed, it must be transported from the nucleus to the cytoplasm to guide protein synthesis. The RNA precursor produced by transcription must be processed to become mature and functional RNA.
Eukaryotic gene transcription and cell cycle
Interphase 1 (G1) – active transcription occurs within the cell
Interphase 2 (G2)—little RNA and protein synthesis
RNA processing and maturation
rRNA processing
The rRNA sequence is transcribed into an RNA precursor, which undergoes processing such as folding, methylation, and terminal modification to form mature rRNA.
Processing of tRNA
Including: shearing and splicing; base modification; 3’-OH linkage-ACC structure
mRNA processing
Generation of 5’ end cap
The eukaryotic cap is formed by a 7-methylguanosine group connected to the 5' end of the mRNA through a 5' end triphosphate bond. The cap structure is not encoded on the DNA, but is added to the first nucleotide of the mRNA before transcription reaches 50 nucleotides.
Main functions: Prevent mRNA from being attacked by phosphatase and nuclease, stabilize the primary structure of mRNA; provide ribosome binding sites, promote the combination of ribosomes and mRNA, and promote protein synthesis.
Generation of 3’ polyA tail
There is a tailing signal sequence (3'-AAUAAA-5') 10-30 nucleotides upstream of the tailing site. The endonuclease recognizes the tailing signal and cuts the mRNA 20 nucleotides downstream of the signal. body, forming a free 3'-OH, and polyA polymerase adds 100-200 adenylate to the 3' end.
Main functions: Increase the stability of mRNA and delay the degradation rate; help transport mature mRNA from the nucleus to the cytoplasm; enhance translation efficiency.
mRNA splicing
Splicing of mRNA: During eukaryotic transcription, both exons and introns are transcribed into hnRNA. hnRNA cuts out the introns under the action of endonuclease, and then connects the various parts of the exons under the action of ligase to become mature mRNA. This process is called mRNA splicing.
Alternative splicing of mRNA: Different tissues or different developmental stages of the same type of cells have different splicing effects, resulting in translation into different protein products, which is called alternative splicing of mRNA.
Processing of micro-RNA
Transcriptome—the spatiotemporal nature of RNA
Transcriptome: The collective name for all RNA after transcription.
The definition includes time and space limitations, and the gene expression of the same cell is not exactly the same in different growth stages and growth environments.
Differences in expression profiles can be used as molecular markers to directly diagnose diseases.
protein biosynthesis
genetic code
Triplet codon - 3 adjacent nucleotides form a triplet codon with no spaces, no overlaps, and no jumps.
Reading frame: The reading frame in which a stretch of nucleotides can be translated into a protein.
Open reading frame: Only one of the three reading frames of a DNA sequence has a coding role and can be translated into the amino acid sequence encoded from the start codon to the stop codon.
Closed reading frame: Some reading frames are blocked due to frequent stop codons and cannot be translated into proteins.
Experiments to decipher the code: Mattei and Nirenberg added artificially synthesized polyU to the in vitro cell-free protein synthesis system and pioneered the method of deciphering the genetic code.
64 triplet combinations, 61 sense codons; 1 start codon; 3 stop codons.
Codon characteristics:
Degeneracy - several codons code for the same amino acid
Universality - With few exceptions, the genetic code of all living things is the same, indicating a common essence and common origin of life.
Preference - Different species and different organisms have different usage frequencies of degenerate codons, which often have species bias.
Mutual recognition of codons and anticodons
Wobble hypothesis: tRNA molecules can be paired and recognized with more than one kind of mRNA codon. The first and second bases of the codon are strictly matched, and the third base is variable.
Ribosome structure and function
The large and small subunits of prokaryotic ribosomes are 50S and 30S
Eukaryotic ribosomes are composed of two subunits, 60S and 40S.
Ribosome functional areas: P site, A site, E site
Protein biosynthesis process: mRNA can combine with multiple ribosomes to simultaneously synthesize multiple peptide chains on one mRNA chain to improve translation efficiency.
Initiation of synthesis: the process in which ribosome large and small subunits, tRNA, and mRNA are combined to form an initiation complex with the assistance of initiation factors
The elongation process of the peptide chain: the information on the mRNA is read from the 5’ end to the 3’ end, and the peptide chain is synthesized from the N end to the C end.
Carry: The second aa-tRNA forms a complex under the action of EF-Tu and GTP and binds to the A site of the ribosome. EF-Tu binds GDP and leaves the ribosome.
Peptide formation: Peptidyl transferase (transpeptidase) catalyzes the fMet carried by the P-position fMet-tRNA to transfer to the A-position to form the first peptide bond with the incoming aa-tRNA. The essence of catalysis is to convert an ester bond into a peptide bond.
Translocation: catalyzed by the translocation factor EF-G, GTP, EF-Ts, and EF-Tu participate. The tRNA at the P position is shed, the ribosome moves along the mRNA, and the tRNA with the peptide chain at the original A position is transferred to the P position, and the next The codon enters the A position for continued translation.
Termination reaction: The release factor RF recognizes the termination codons UAA, UAG, and UGA that enter the A position of the ribosome. The peptidyltransferase on the large subunit allosterizes, showing the activity of the hydrolase, so that the polypeptide chain carried by the tRNA on the P position is separated from the tRNA. The ester bond between them is hydrolyzed. The tRNA falls off from the P position, and the 70S ribosome immediately falls off from the mRNA, dissociates into the 30S and 50S subunits, and is put into the next round of ribosome cycle to synthesize another new protein molecule.
Characteristics of eukaryotic protein synthesis
The starting tRNA of eukaryotes is Met-tRNA and does not require N-terminal formylation; the starting tRNA of prokaryotes is fMet-tRNA.
The eukaryotic 40S small subunit first binds to Met-tRNA and then to mRNA; the prokaryotic small subunit first binds to mRNA and then to fMet-tRNA
The main sign of recognition between eukaryotic 40S small subunit and mRNA is the cap structure; prokaryotic mRNA recognizes SD sequence
post-translation processing
Excision of methionine or formylmethionine at the N-terminal end of the peptide chain; chemical modification (methylation, acetylation) of amino acid residues in the peptide chain; formation of disulfide bonds; excision of the signal peptide; removal of the peptide chain Folding; excision of functionally unnecessary peptide segments from the precursor
Cleavage and processing of peptides
chemical modification of peptides
Phosphorylation refers to the process of transferring the phosphate group of ATP or GTP to the amino acid residue of protein, catalyzed by protein kinase, and plays an important role in the process of cell signal transduction. Phosphorylation and dephosphorylation are key to controlling the cell cycle. Serine phosphorylation can often improve protein activity; tyrosine phosphorylation mainly promotes interactions between proteins and forms multi-protein complexes.
Glycosylation is the process of attaching sugars to proteins or lipids under the control of enzymes, and occurs in the endoplasmic reticulum or Golgi apparatus. Proteins undergo glycosylation to form glycoproteins. The protein structure changes, resists the degradation effect of proteases, improves the solubility of the protein, and allows the protein to accurately enter the respective organelles. Some proteoglycans are secreted outside cells to form extracellular matrix or mucus layer, and some are anchored on membranes and have a protective effect on cells.
Acetylation refers to the process of adding an acetyl group to a protein lysine residue under the action of acetyltransferase. It is a mechanism by which cells control gene expression, protein activity or physiological processes. In the nucleus, the processes of histone acetylation and histone deacetylation are in dynamic balance, accurately regulating gene transcription and expression. The acetylation of histone lysine residues causes the side chain to no longer be positively charged and loses the ability to bind tightly to DNA, which is conducive to the detachment of DNA from nucleosomes. The degree of acetylation of core histones represents the expression of gene transcription activity. High and low.
Secreted protein: A protein that is secreted outside the cell after being synthesized inside the cell. For example: salivary amylase, pepsin, digestive enzymes, antibodies and some hormones.
Secreted proteins synthesized on ribosomes pass through the endoplasmic reticulum and Golgi apparatus rather than being transported directly to the cell membrane.
Signal peptide: a hydrophobic amino acid sequence at the beginning of the peptide chain, which is recognized and combined with receptors on the endoplasmic reticulum membrane. It reaches the lumen of the endoplasmic reticulum through the protein pores in the membrane, and is then recognized by the signal peptide located on the lumen surface. Enzymatic hydrolysis and fragmentation.
Signal peptide sequence: Starting from the start codon, a sequence encoding a signal peptide.
Protein synthesis and cell cycle
As cells prepare to enter division, protein synthesis accelerates; genome replicates
When cells are in mitosis, protein synthesis is inhibited
Proteome: All types of proteins expressed by a cell under specific physiological or pathological conditions are called proteome.
The definition of proteome includes time and space limitations, and the types of proteins expressed by different cells in different physiological or pathological states are also different.
Gene expression and regulation
Eukaryotic gene expression regulation
DNA level
gene loss
During the cell differentiation process of lower eukaryotes, some somatic cells can achieve the purpose of gene regulation by losing certain genes to remove the activity of these genes. This process is irreversible.
gene amplification
The copy number of certain specific genes in cells increases specifically and massively, allowing cells to produce enough gene products to meet certain needs in a short period of time.
gene rearrangement
Changes in the structure of the genome change the way cells synthesize protein types.
The impact of chromatin structure on transcriptional regulation
DNA methylation, histone acetylation
RNA levels - eukaryotic regulation of transcription initiation activity
Cis-regulatory element: a component of a gene that controls the start time and degree of gene expression (transcription), recognizes each other with transcription factors (TF), and controls gene activity, including promoters, enhancers, silencers, etc. Adjustment element
Enhancer: There is a core characteristic sequence, which is independent of directionality, position, and distance. It is tissue-specific and requires a promoter to function.
trans element
Transcription factors: proteins encoded by specific genes, generally with a relatively special structure.
Molecular domains of transcription factors: DNA binding domain, dimerization domain and transcription activation domain
RNA level - regulation at the post-transcriptional level in eukaryotes
Alternative splicing of pre-mRNA: refers to the process of producing different mRAN splicing isoforms from one pre-mRNA through different splicing methods.
RNA editing: After transcription, mRNA changes the original information of the DNA template by inserting, deleting, or replacing bases, thereby expressing proteins with a variety of different amino acid sequences.
Long noncoding RNAs regulate transcription and translation
Protein level - translation level
Stability of mRNA: The lifespan of mRNA is affected by its own structure and other factors within the cell. The 5' cap and 3' tail help increase the stability of the mRNA molecule.
Regulation of translation initiation efficiency by mRNA structure
The 5’ untranslated region is involved in regulating the initiation of translation
The position of the start codon and its flanking sequences affect translation efficiency
The structure of 3’-UTR regulates translation
The effect of miRNA on target mRNA and whether the sequence is complementary
miRNA regulation
Incomplete pairing and binding of miRNA and target mRNA mainly affects the translation process
The miRNA completely pairs with the target mRNA, causing specific cleavage and degradation of the target mRNA.
miRNA is conserved among species, and its expression is temporal and tissue-specific.
Regulation of translation by antisense RNA (RNA molecules complementary to mRNA)
application
Antisense RNA regulates target gene expression by inhibiting translation templates
Used for gene function verification; inhibition of animal and plant viruses; gene therapy, etc.
Protein level - post-translational processing and transport
splicing of peptide chain
Amino acid chemical modification
folding of polypeptide chain
Targeted delivery of proteins
Prokaryotic gene expression regulation
In order to adapt the metabolic process to changes in the environment and maintain their own survival and reproduction, prokaryotes have a set of mechanisms to accurately regulate gene expression and protein synthesis.
Operator models (Jacob and Monod)
Operon: Found in prokaryotes, regulatory genes, operator genes and structural genes are arranged in clusters and together form a transcriptional functional unit. The expression of structural genes is controlled by a shared regulatory region.
Specific example - Glucose lactose sensitivity operon
Structure of the lactose operon
Three structural genes encoding enzymes involved in lactose catabolism
Upstream of the structural gene, from near to far, are the operator gene lacO, promoter PZYA, regulatory gene lacI and regulatory gene promoter PI
There is also a binding site for metabolite activator protein (CAP) upstream of the promoter PZYA.
Negative regulatory mechanism of lactose operon
Without a lactose inducer, the repressor protein binds to the operator gene, preventing RNA polymerase from binding to the promoter. The structural gene is not transcribed, inhibiting lactose catabolism.
Lactose or its derivatives bind to the repressor protein, causing it to undergo a conformational change and lose the ability to bind to the operator gene. RNA polymerase initiates the normal transcription of the structural gene, encoding lactose decomposing enzyme, which decomposes lactose into galactose and glucose.
Negative feedback: After lactose is decomposed into galactose and glucose, the lack of lactose molecules causes the repressor protein to bind to the operator gene, shutting down the expression of structural genes.
Positive regulatory mechanism of lactose operon
In the absence of glucose, the lactose operon transcriptional activity is enhanced. Adenylyl cyclase converts ATP into cyclic adenosine monophosphate (cAMP). cAMP combines with CAP to form a dimer, and then combines with a specific DNA sequence (CAP binding site) to increase the lactose operon transcription efficiency. 50 times.
In the presence of glucose, the cAMP concentration decreases and the binding of cAMP to CAP is blocked, so the expression of the lactose operon decreases.
When glucose is available, E. coli preferentially utilizes glucose
The lactose operon regulates the carbon source utilization process in Escherichia coli
The lactose operon is an automatic control system for regulating lactose metabolism in Escherichia coli
When glucose is present, bacteria preferentially choose glucose to supply energy. Glucose inhibits the transcription of the lactose operon by reducing the concentration of cAMP and preventing the binding of cAMP to CAP, so that bacteria can only utilize glucose.
In the absence of glucose but only lactose, the repressor protein depolymerizes the lacO sequence. At the same time, CAP binds to cAMP and acts on the CAP site of the lactose operon to activate transcription, allowing the bacteria to efficiently utilize lactose and decompose lactose into galactose and glucose. ,provide energy
CAP is a positive regulatory factor and lactose repressor protein is a negative regulatory factor
Specific example - Attenuation of the tryptophan operon
Structure of the tryptophan operon
Regulatory gene trpR, promoter P, operator gene trpO, leader sequence trpL and attenuator trpa, as well as structural gene trpEDCBA - tryptophan synthesis gene. Regulating genes away from operons
Leader sequence upstream of coding genes: weak rho factor-independent transcription terminator
Attenuation effect:
When tryptophan is at a high level, the ribosome translates the leader peptide and stops at the stop codon. The ribosome covers regions 1 and 2, and regions 3-4 of the leader sequence form a pair, and RNA polymerase terminates transcription.
When tryptophan is starved, the ribosome pauses at two tryptophan codons. The ribosome occupies region 1, and regions 2-3 of the leader sequence form a pair. RNA polymerase continues to move forward, and structural genes are transcribed.
Timing control
changes in genetic material
Gene mutation
Detectable and heritable changes in genetic material at the genetic level
General characteristics of genetic mutations
Reproducibility of mutations—recurrence of the same species in different individuals, at different times, and in different places
Reversibility of mutations - positive mutations, reverse mutations
Mutation pleiotropy - multiple alleles
Parallelism of mutations - similar mutations occurring in similar species
Low frequency of mutations - mutations occur less frequently within a certain period of time
Advantages and disadvantages of mutations - diversity and selection
Molecular basis of genetic mutations
Type of genetic mutation
Base substitution: The phenomenon in which one base pair is replaced by another base pair in a DNA molecule, including transitions and transversions.
Indel: The introduction or loss of 1 or more bases.
Genetic effects of base substitutions and frameshift mutations
Genetic effects of mutations in coding regions
Synonymous mutation: After base substitution, the new codon generated on the mRNA represents the same amino acid as the original codon, without causing changes in the protein sequence.
Missense mutation: a base substitution in the DNA sequence that changes the codons on the mRNA, resulting in amino acid substitutions and the genetic effects of mutations in the non-coding region.
Nonsense mutation: After base substitution, nonsense codons in the mRNA appear early, resulting in an incomplete peptide chain.
Frameshift mutation: The addition or deletion of base pairs in the genome changes the reading frame of the gene, causing the codons after the site to change.
Genetic effects of mutations in non-coding regions
The mechanism by which genetic variation occurs
Chemically induced mutations
Induced mutations of base analogs: During the replication process, base analogs are incorporated into DNA, and their tautomers pair with bases in different ways, causing base substitutions
Chemical mutagens that change the chemical structure of DNA: Some nitrites, alkylating agents and hydroxylamine can change the chemical structure of nucleotides in DNA, thus leading to base substitutions.
Mutagenic compounds that bind to DNA molecules
High-energy ray-induced mutations: ultraviolet rays
Recombination: The rearrangement of chromosomes is recombination; recombination occurs during meiosis and also occurs in somatic cells of eukaryotes.
homologous recombination
Under the action of recombinase, any pair of homologous sequences in two DNA molecules serve as substrates for exchange.
site-specific recombination
It does not rely on the homology of DNA sequences, but on specific DNA sequences that can bind to certain enzymes that can catalyze the breakage and rejoining of DNA strands.
transposition recombination
Transposition means that a DNA sequence can be copied or broken from the original site, circularized and inserted into another site. If the site is internal to the gene, the transposed fragment often causes frameshift mutations or gene inactivation.
Transposable elements are divided into three categories: cut-and-paste transposons, replicating transposons, and retrotransposons
Suppression of mutations and repair of DNA damage
mutation suppression
Codon mergers - synonymous mutations (main method)
Suppression of intragenic mutations—double frameshift mutations
Suppression of intergenic mutations: suppression of nonsense mutations and missense mutations, suppression of frameshift mutations
DNA polymerase replication repair
DNA mismatch repair system
Repair of pyrimidine dimers - photorepair
Excision repair - dark repair
Reorganization
DNA polymorphism
basic concept
Genetic polymorphism: In a population, the phenomenon of two or more alleles of a gene locus.
DNA polymorphism: In a population, there are two or more variant types at one sequence site.
Genetic markers: refer to material markers that can be used to distinguish biological individuals or groups and their specific genotypes and can be stably inherited.
Genetic marker type
Phenotypic markers—appearance, behavior, and other characteristics
Protein markers - differences in protein molecular size and configuration
DNA markers – differences in base composition
Single Nucleotide Polymorphism (SNP)
Small insertion and deletion polymorphisms (indels)
Number of tandem repeats polymorphism (VNTR)
copy number variation (CNP)
Detection and Application of Genetic Markers
PCR and enzyme digestion combined to detect SNP
PCR and sequencing combined to detect microsatellite polymorphisms
PCR and electrophoresis combined detectioninde
DNA chip
Characteristics of an ideal genetic marker
Number of markers - single, small amount, abundant (sequencing, chip)
Marker distribution - concentrated, dispersed (SNP)
Marker discrimination - dominance, co-dominance (differentiating heterozygotes)
Genetic polymorphism – dimorphism, trimorphism or higher (fingerprint)
Application of DNA polymorphism markers in animal genetic breeding
Genomes and Genetic Engineering
Genome overview
Genome structural features
Genome: The collection of all DNA carried by a set of chromatids of a species.
Category 1
Single sequence: one or a few copies of a DNA sequence (50-80% of the mammalian genome)
Repeating sequences: sequences that appear hundreds or even thousands of times
Category 2
Highly repetitive sequences: high repetition frequency - inverted repeats, tandem repeats, interspersed repeats
Inverted repeat sequence: palindrome or mirror repeat
Tandem repeats: satellite DNA (centromeres), minisatellite DNA (telomeres), and microsatellite DNA
Interspersed repetitive sequences: transposons, etc.
Moderately repetitive sequences: repeated dozens to tens of thousands of times - long scattered fragments, short scattered fragments
Long scattered fragments (LINE): KpnI family, 3-5kb in length, scattered distribution
Short scattered fragments (SINE): Alu sequence, length 300bp, average 5kb Alu sequence
Multigene family: A group of genes with similar sequences that encode several genes of a protein family that are structurally and functionally related.
Pseudogene: In the same multigene family, some members are similar to other genes but do not produce functional gene products.
Genome C value and C value paradox
C value refers to the total amount of haploid genomic DNA in each organism
The maximum C value refers to the total amount of haploid genomic DNA in each organism.
The minimum C value refers to the total content of DNA encoding genetic information (ie, exonic DNA) in each organism.
C-value paradox and related explanations
C-value paradox: There is no strict correspondence between the C-value of a species and its evolutionary complexity, that is, the complexity of organisms does not increase entirely in proportion to the size of the genome.
Possible reasons for the C-value paradox:
Variation in genome size (C value) is primarily caused by variation in non-coding DNA segments
There are different numbers of repeat sequences on chromosomes, resulting in a large range of changes in genome C values.
The Origin and Evolution of Genomes—The “RNA World” Theory
The transformation from “RNA world” to “DNA” world:
The coding function of RNA is replaced by DNA, which is more stable
The catalytic function of RNA is replaced by proteins, which are more diverse
RNA still retains its regulatory function, but RNA is more flexible
The evolution of the genome
generation of new genes
genetic map
A map used to describe the distribution status of genes on chromosomes, linear arrangement and other information
Genome map type
genetic map
The linear arrangement of genes or markers on chromosomes obtained through genetic recombination analysis is called a genetic linkage map. It calculates the recombination frequency between linked genetic markers and determines their relative distance, which is generally expressed in centimorgans (cM, that is, the recombination frequency of each meiosis is 1%).
physical map
The linear arrangement of genes or markers on chromosomes determined through restriction enzyme fragment arrangement, or using hybridization, sequencing and other techniques is called a physical map. It is often expressed as the physical distance between genetic markers (bp, i.e. the number of base pairs, or kb/Mb, etc.).
Construction of linkage map
Linkage analysis: Locate the relative positional relationship between genetic markers or genes based on the recombination rate of two adjacent genetic markers or genes located on the same chromosome.
Physical mapping: Use methods such as chromosome in situ hybridization or somatic cell hybridization to locate genes or genetic markers on a certain chromosome or a specific region of a chromosome.
Typical physical maps include:
chromosome map
fluorescence in situ hybridization
Restriction map
Complete genome sequence
The significance of genome analysis
Analysis of key genes controlling complex traits
Molecular genetic testing
Marker assisted selection
Whole genome selective breeding
Excavation of excellent genetic germplasm resources
Overview of Animal Genetic Engineering
Definition: Based on the theoretical basis of molecular genetics and using modern methods of molecular biology and microbiology as a means, genes from different sources are constructed in vitro according to a pre-designed blueprint, and then introduced into living cells to change the biological origin. certain genetic characteristics, obtain new varieties, and produce new products. Genetic engineering technology provides a powerful means for studying gene structure and function.
Main operating techniques of genetic engineering
Obtain DNA fragments that meet the requirements - amplification, enzyme digestion, artificial synthesis
Construct a gene expression vector - use DNA ligase to connect the vector and foreign fragments
Introduce the gene of interest into the recipient cells
Detection and identification of target genes
Animal transgenic technology applications and examples
human disease models
Bioreactor for pharmaceutical proteins
Discover and verify gene function
Cultivation and improvement of animal breeds
Animal gene editing technology applications and examples
Gene editing technology: refers to editing the target position of the genome to achieve the knockout, addition or rewriting of specific DNA fragments.
Bacterial CRISPR/Cas defense system against viruses
How the CRISPR Cas9 system works