Sample Submission Guidelines
Turn In Vivo Delivery Questions into Sequencing-Supported Evidence
In vivo gene therapy studies often begin with a direct question: where did the delivered genetic material go? Once the project includes multiple tissues, groups, or timepoints, that question becomes more complex.
You may need to know whether a vector is detectable in target tissues, whether non-target tissues show signal, whether the intended sequence is supported by reads, whether RNA-level expression is present, or whether integration-related analysis should be considered for your vector system.
We help you turn those questions into a sequencing and biodistribution plan that fits the study, instead of forcing every project into the same assay.
What this solution helps you answer
- Is the delivered vector or genetic material detected in selected tissues?
- How do target and non-target tissues compare across groups?
- Is the vector genome or target region supported by sequencing reads?
- Does the transgene show RNA-level expression in selected tissues?
- Should integration-site or insertional event assessment be added?
- What QC and bioinformatics outputs are needed for internal review?
The goal is to choose the right readouts before samples are processed, so the final data package answers the question you actually need to ask.
A tissue-level biodistribution readout may show whether a target sequence is detectable. It may not confirm the broader vector sequence, explain RNA-level expression, or address integration-related questions.
That is why some in vivo gene therapy projects need a combined approach:
- Biodistribution readouts for tissue-level presence or copy-related evidence
- Targeted sequencing for vector or target-region confirmation
- AAV genome sequencing for vector genome structure and sequence support
- RNA sequencing for transgene expression or host response profiling
- Whole-genome or long-read sequencing for broader genomic context
- Integration-site analysis when vector biology and study design require it
We help you decide which layers are necessary, which are optional, and which may not be useful for the current study.
Our Service Capabilities for Gene Therapy Biodistribution Projects
We support in vivo gene therapy sequencing projects as integrated study packages, not isolated data-generation tasks. Before the project starts, our team can review sample type, vector design, readout goals, sequencing strategy, QC checkpoints, and expected deliverables with you.
That early review matters. It helps prevent a common problem in complex studies: generating data that are technically valid but not well matched to the biological question.
Vector and Delivery Systems We Can Support
- AAV vector research
- Lentiviral or retroviral vector research
- Plasmid or non-viral delivery studies
- LNP or nanoparticle-delivered nucleic acid studies
- Engineered cell or exogenous gene delivery contexts
Sequencing and Analysis Modules
- Vector genome or target-region confirmation
- Tissue biodistribution readouts
- Transgene expression profiling
- Integration-site or insertional event assessment where applicable
- Multi-tissue or multi-timepoint comparison
- Bioinformatics reporting and visualization
Project Execution Support
A strong project starts before sequencing. We help you review sample type and grouping, tissue list and timepoints, construct map or target sequence availability, DNA or RNA QC information, sample naming and labeling, intended output tables and figures, and required bioinformatics file types.
This helps your team scope the project more clearly and reduces the risk of collecting data that cannot answer the original study question.
Choose the Right Readout: Biodistribution, Sequence Confirmation, Expression, or Integration
A good in vivo gene therapy study does not start with a platform. It starts with the question your team needs to answer. The table below shows how common readouts differ and when each one may fit.
| Readout / Method | Main Question Answered | Best-Fit Use Case | Common Sample Type | Typical Outputs | Important Limitation |
|---|---|---|---|---|---|
| qPCR / ddPCR-style quantification | Is the target sequence detectable or quantifiable? | Tissue biodistribution, vector copy-related analysis, known target detection | Extracted DNA or RNA, tissue-derived nucleic acid | Copy-related tables, tissue-level comparison, normalized signal output | Limited sequence context; does not confirm broad vector structure |
| Targeted Region Sequencing | Is the expected vector or target region supported by reads? | Known target confirmation, amplicon-based vector sequence support | DNA, amplicons, prepared target regions | Read support, coverage summary, target-region table | Focused on selected regions; not designed for genome-wide discovery |
| AAV Genome Sequencing | Is the AAV genome structure or sequence supported? | AAV vector sequence confirmation and vector-related QC | Vector DNA or prepared AAV-related material | Vector sequence evidence, coverage, sequence structure support | Vector-focused; does not replace tissue-level biodistribution analysis |
| RNA Sequencing | Is the transgene expressed, and is host response relevant? | Transgene expression profiling, tissue response, pathway-level analysis | Total RNA from tissue or cells | Expression matrix, heatmap, differential expression table, pathway output | RNA signal does not always equal vector DNA presence |
| Whole Genome Sequencing | Is broader genomic context needed? | Genome-wide variant or context analysis, selected research questions requiring genomic context | Genomic DNA | Variant tables, genome-wide data, QC summary | More complex and not always necessary for focused biodistribution |
| Long-Read Sequencing | Is longer-range sequence context important? | Structural context, larger insert-related questions, selected vector or integration studies | High-quality DNA or prepared material | Long-read alignment, structural context, candidate region support | Requires higher-quality input and careful study design |
| Integration Site Analysis | Where are candidate insertion events detected? | Lentiviral, retroviral, transposon, or other integration-relevant systems | Genomic DNA | Candidate insertion table, genomic location summary, visualization | Only appropriate when the vector biology and study question justify it |
For related service details, see our AAV Genome Sequencing, Targeted Region Sequencing, Whole Genome Sequencing, and Lentiviral/Retroviral Integration Sites Analysis pages.
If the main question is where the genetic material is detected, start with a biodistribution readout. If the main question is whether the expected vector or target region is present and supported by reads, add targeted sequencing or vector genome sequencing. If the main question is whether the transgene is expressed, add RNA-level analysis.
If the main question is whether insertion-related events matter, consider integration-site analysis only when the vector type and study design make it relevant. If your team needs a clear internal data package, we should define the bioinformatics deliverables before samples are submitted. That includes the expected tables, figures, file types, QC summaries, and report notes.
Sample-to-Report Workflow with QC Checkpoints
Our workflow combines technical processing with project management. Once samples enter the project, we track whether each step still supports the original study question.
Step 1: Project Intake and Readout Selection
Before samples enter the workflow, we review the study design with your team. Useful information includes vector or delivery system, species and tissue list, target and non-target tissues, group design and timepoints, sample format, target sequence or construct map, desired readouts, and expected output tables and figures.
This step helps us decide whether your study needs one readout or a combined package.
Step 2: Sample Submission and Pre-QC Review
When samples are prepared for shipment, clear labeling and documentation matter. CD Genomics asks customers to provide a completed sample submission form, keep sample names consistent between the form and tube labels, and submit electronic QC data when available.
For tube submission, 1.5 mL centrifuge tubes are often suitable. Plates should be sealed tightly to reduce sample loss or cross-contamination. DNA in water or TE buffer should be shipped with ice packs, while RNA, cells, bacteria, and frozen tissues should be shipped with dry ice.
At this stage, our team checks sample identity, sample type, labeling, and available QC information. If something is unclear, we clarify it before the sample moves further into processing.
Step 3: Nucleic Acid Extraction and Quality Control
The technical workflow depends on the input material and the selected readout.
For DNA-based readouts, extracted DNA should be RNase-treated and show no obvious degradation or contamination. CD Genomics sample guidance recommends DNA OD260/280 as close to 1.8-2.0 as possible.
For RNA-based readouts, total RNA should be DNA-free. CD Genomics guidance recommends A260/A280 ≥ 1.8, A260/230 ≥ 1.8, and RIN ≥ 6 for total RNA samples.
QC review may include concentration, purity, degradation, and suitability for the selected sequencing method. If the sample quality does not match the planned readout, we discuss possible adjustments before proceeding.
Step 4: Library or Assay Preparation and Sequencing
After QC, the project moves into the selected technical route.
For targeted sequencing, target regions are amplified or enriched before sequencing. For AAV genome sequencing, vector-related sequence support and coverage are assessed. For RNA sequencing, RNA is converted into sequencing-ready libraries. For whole-genome or long-read strategies, library preparation depends on DNA quality and the required data type.
The technical aim is not only to generate reads. It is to generate data that match the planned biological readout.
Step 5: Bioinformatics, QC Summary, and Report Delivery
After sequencing, we process the data into usable outputs. Depending on the project, this may include raw data and clean data, sample-level QC summary, target-region read support, tissue-level distribution tables, expression matrix, integration-site candidate table where applicable, visual summaries, analysis report notes, and pipeline and parameter records where included.
The final package is built to help your team review the study, compare groups, and decide whether follow-up experiments are needed.
Sample Requirements for Multi-Tissue Gene Therapy Studies
Sample needs depend on vector type, readout, species, tissue, and extraction status. The table below gives practical starting points based on CD Genomics sample guidance and common sequencing project planning. Final requirements should be confirmed during project review.
| Sample Type | Recommended Input | Container | Shipping | QC Checkpoints | Notes |
|---|---|---|---|---|---|
| Frozen tissue for DNA/RNA extraction | Project-specific; for RNA-seq, tissue guidance may start from 10-50 mg depending on application | Cryotube or approved sealed tube | Dry ice | Tissue identity, degradation risk, DNA/RNA suitability | Provide tissue name, group, timepoint, vector type |
| Extracted genomic DNA for short-read sequencing | WGS: ≥500 ng recommended; WES: ≥500 ng recommended; viral genome sequencing: ≥1 μg recommended | DNase-free tube | Ice packs | OD260/280 close to 1.8-2.0, concentration, degradation, contamination | Submit construct map or target sequence if available |
| Extracted genomic DNA for long-read sequencing | PacBio WGS: ≥3 μg; Nanopore WGS: ≥5 μg | DNase-free tube | Ice packs | High molecular weight DNA, concentration, purity | Useful when longer-range sequence context is needed |
| Extracted RNA for expression profiling | Whole transcriptome: ≥3 μg recommended; mRNA-seq: ≥500 ng recommended | RNase-free tube | Dry ice | A260/A280 ≥1.8, A260/230 ≥1.8, RIN ≥6 | Useful for transgene expression or host response profiling |
| Cells for RNA-based work | mRNA-seq guidance may start from ≥1×106 cells | Approved tube or frozen cell format | Dry ice | Cell amount, RNA integrity after extraction | Provide cell type, group, treatment, and target readout |
| Blood sample | For selected assays, 0.5-1 mL or more may be required depending on readout | Plastic anticoagulant blood collection vessel | Rigid protected packaging; condition depends on assay | Sample integrity and matrix suitability | Provide anticoagulant type and study group |
| Purified amplicon | Amplicon sequencing: ≥1 μg recommended; 500 ng minimum | Low-bind tube | Ice packs | Concentration, single target band if available | Useful for focused target-region confirmation |
| Viral particles or vector material | Project-specific feasibility review required | Approved frozen format | Dry ice | Identity, concentration if available, construct information | Useful for vector-related sequence confirmation |
Bioinformatics Analysis and Deliverables
Bioinformatics should be planned before sequencing begins. When we understand your vector type, target region, tissue list, and reporting goals early, we can build a more useful output package.
Minimum Deliverables
- Raw data and clean data where applicable
- Sample-level QC summary
- Target-region or vector-related read summary
- Tissue-level distribution table
- Basic visualization files
- Method and analysis notes
- Final report package
For projects involving expression profiling, outputs may also include expression matrices and heatmaps. For projects involving integration-related questions, outputs may include candidate insertion tables and genomic location summaries.
Optional Add-ons
- Integration-site candidate analysis
- RNA expression matrix and differential expression
- Host response pathway analysis
- Multi-timepoint or multi-dose comparison
- Custom figure-ready visualization
- Pipeline parameter record
- Cross-readout summary table
For RNA-level projects, see our RNA Sequencing service. For customized analysis support, see our Bioinformatics services.
Application Scenarios
These study scenarios help show how sequencing and biodistribution readouts can be combined when the research question requires more than one layer of evidence.
AAV Tissue Tropism and Vector Biodistribution
AAV studies often need to compare target tissues with non-target tissues after in vivo administration. Depending on the goal, the project may combine biodistribution readouts with AAV Integration Site Analysis or AAV genome-related sequencing.
Lentiviral or Integrating Vector Insertion-Site Assessment
For lentiviral, retroviral, transposon, or other integration-relevant systems, integration-site analysis may be needed to understand candidate insertion locations. This analysis should be considered only when the vector biology and study design justify it.
Non-Viral Delivery and Nucleic Acid Persistence Studies
For plasmid, LNP, or nanoparticle-based delivery systems, the study may focus on where nucleic acid material is detected, whether target regions are supported by sequencing, and whether expression readouts are needed.
Transgene Expression and Tissue-Level Response Profiling
When vector presence alone is not enough, RNA-level analysis can help show whether transgene expression or tissue-level response is part of the data package.
References
- Development and validation of a model gene therapy biodistribution assay for AVGN7 using digital droplet polymerase chain reaction
- Adeno-associated virus as a delivery vector for gene therapy of human diseases
- Characterization of AAV vectors: A review of analytical techniques and critical quality attributes
- A novel approach to quantitate biodistribution and transduction of adeno-associated virus gene therapy using radiolabeled AAV vectors in mice
- Mapping administration route-dependent transduction profiles of commonly used AAV variants in mice by barcode amplicon sequencing
- Auto-expansion of in vivo HDAd-transduced hematopoietic stem cells by constitutive expression of tHMGA2
Demo Results: What Your Data Package May Look Like
Demo results help your team understand what the final outputs may look like. The examples below are common result formats that can be included when they match the study design.
Demo 1: Tissue Biodistribution Profile
A tissue biodistribution profile can show how vector-related signal is distributed across selected tissues, groups, or timepoints. A typical table or chart may include sample ID, tissue type, group or dose, timepoint, target signal, normalized output, and QC status. This view helps your team compare target and non-target tissues in one place.
Demo 2: Vector Sequence and Target-Region Confirmation
A sequence confirmation output can show whether reads support the expected vector region, transgene region, or selected target sequence. A typical report view may include target region, read count or read support, coverage summary, sequence match notes, variant or mismatch flags where relevant, and QC interpretation notes.
Demo 3: Expression and Integration-Related Outputs
For projects that include RNA or integration-related questions, a combined output may show expression patterns and candidate insertion information. A typical output may include tissue or sample group, transgene expression level, host response markers if included, candidate insertion location where applicable, genomic feature annotation, supporting read information, and QC comments.
FAQ: Planning an In Vivo Gene Therapy Sequencing Project
1. Which readout should I choose first?
Start with the question you need to answer. If you need tissue-level presence, start with biodistribution. If you need sequence support, add targeted or vector sequencing. If you need expression evidence, add RNA-level analysis. If insertion-related events matter, integration-site analysis may be useful.
2. Can tissue, blood, DNA, and RNA samples be included in one project?
Yes, but they may require different handling, QC checks, and sequencing workflows. We review the sample types first, then align each sample group with the correct readout.
3. When is sequencing more useful than qPCR or ddPCR alone?
Sequencing is useful when copy-related detection is not enough. It can add target-region support, vector sequence information, expression profiling, integration-related evidence, or broader genomic context.
4. Can you help with AAV genome confirmation?
Yes. For AAV-related work, we can help assess whether AAV genome sequencing or target-region sequencing fits the project. The best option depends on the vector design, sample type, and question being asked.
5. When should integration-site analysis be considered?
Consider it when the delivery system can integrate, when insertion-related questions are part of the study, or when genomic location information is needed. It is not necessary for every biodistribution project.
6. What information should I send before requesting a project plan?
Useful information includes vector type, species, tissue list, timepoints, sample format, target sequence, construct map, desired readouts, and any existing DNA or RNA QC data.
7. What bioinformatics outputs can be included?
Depending on the project, outputs may include QC summaries, tissue-level tables, target-region read support, expression matrices, candidate insertion tables, visualizations, and report notes.
8. Can this solution support multi-tissue, multi-timepoint designs?
Yes. Multi-tissue and multi-timepoint designs are often a good fit for this solution when sample quality, grouping, and readout goals are planned before sequencing begins.
Customer Publication Case: Sequencing Evidence in an In Vivo Gene Therapy Study
Customer Publication Case
Journal: Molecular Therapy: Methods & Clinical Development
Published: 2024
DOI: 10.1016/j.omtm.2024.101319
Background
This customer publication studied an in vivo hematopoietic stem cell gene therapy approach designed to avoid ex vivo cell harvesting, manipulation, and transplantation. The authors constructed HSC-tropic HDAd5/35++ vectors expressing a truncated HMGA2 gene and a GFP reporter gene. A SB100x transposase vector mediated random integration of the transgene cassette.
The study is relevant here because it shows why in vivo gene therapy research may need more than one type of sequencing-supported evidence. The authors examined cell expansion, lineage involvement, insertion-site behavior, genomic stability, and validation in humanized mice.
Methods
The study mobilized HSCs in mice using G-CSF and AMD3100/Plerixafor, followed by intravenous injection of an integrating tHMGA2/GFP vector. The authors followed GFP-positive peripheral blood mononuclear cells over time and evaluated expansion across HSCs and myeloid, lymphoid, and erythroid progenitor populations in bone marrow and spleen.
The sequencing-related methods included genome-wide integration site analysis and whole-exome sequencing. These methods helped the authors examine whether expansion was polyclonal and whether broader genomic instability indices differed between treated and control mice.
Results
The paper reports that GFP-positive peripheral blood mononuclear cells showed slow but progressive expansion, reaching about 50% by week 44. Expansion was also observed in secondary recipients. The authors reported expansion at the HSC level and across progenitor populations in bone marrow and spleen.
Figure 5 presents genome-wide insertion site analysis by TRACE sequencing. The figure includes unique insertion-site-containing DNA fragments, unique genomic insertion coordinates, sequence logos around insertion sites, gene-region distribution, and chromosome-level insertion location visualization.
A key observation from the study was that expansion was polyclonal, with no signs of clonal dominance based on genome-wide integration site analysis. Whole-exome sequencing did not show significant differences in genomic instability indices between tHMGA2/GFP mice and untreated control mice.
Figure 5 from the customer publication shows genome-wide insertion site analysis in an in vivo HDAd-transduced HSC gene therapy study, helping readers understand how insertion-site information can support a broader sequencing evidence package.
Conclusion
This customer publication shows why a multi-readout sequencing plan can be useful in in vivo gene therapy research. Depending on the vector system and study goal, a project may need tissue-level evidence, sequence-level confirmation, insertion-site information, expression data, and broader genomic context.
For a service project, the practical lesson is simple: define the question first, then build the sequencing and bioinformatics plan around that question.
Reference
Related Customer Publications
The publications below are relevant to gene therapy, vector delivery, sequencing, or immune-response research contexts. They are listed as related customer publications, not as full case studies.
| Publication | Journal / Year | Related Service Tag | Why It Is Relevant |
|---|---|---|---|
| Auto-expansion of in vivo HDAd-transduced hematopoietic stem cells by constitutive expression of tHMGA2 | Molecular Therapy: Methods & Clinical Development, 2024 | Whole Exome Sequencing, WES | Closest fit for the customer publication case; relevant to in vivo gene therapy and genomic assessment |
| In vivo base editing rescues ADPKD in a humanized mouse model | Nature Communications, 2025 | RNA-seq / RNA-seq Library Construction and Sequencing | Relevant to in vivo AAV-delivered base editing and transcriptome-wide RNA editing analysis |
| The HLA class I immunopeptidomes of AAV capsid proteins | Frontiers in Immunology, 2023 | HLA Typing | Relevant to AAV capsid immunopeptidome research and gene delivery immune-response context |
See more articles published by our clients.
